Neutral Lipid Storage Disease Research Articles

CGI-58 facilitates lipolysis on lipid droplets but is not involved in the vesiculation of lipid droplets caused by hormonal stimulation

A lipid droplet (LD)-associated protein, perilipin, is a critical regulator of lipolysis in adipocytes. We previously showed that Comparative Gene Identification-58 (CGI-58), a product of the causal gene of Chanarin-Dorfman syndrome, interacts with perilipin on LDs. In this study, we investigated the function of CGI-58 using RNA interference. Notably, CGI-58 knockdown caused an abnormal accumulation of LDs in both 3T3-L1 preadipocytes and Hepa1 hepatoma cells. CGI-58 knockdown did not influence the differentiation of 3T3-L1 adipocytes but reduced the activity of both basal and cAMP-dependent protein kinase-stimulated lipolysis. In vitro studies showed that CGI-58 itself does not have lipase/esterase activity, but it enhanced the activity of adipose triglyceride lipase. Upon lipolytic stimulation, endogenous CGI-58 was rapidly dispersed from LDs into the cytosol along with small particulate structures. This shift in localization depends on the phosphorylation of perilipin, because phosphorylated perilipin lost the ability to bind CGI-58. During lipolytic activation, LDs in adipocytes vesiculate into micro-LDs. Using coherent anti-Stokes Raman scattering microscopy, we pursued the formation of micro-LDs in single cells, which seemed to occur in cytoplasmic regions distant from the large central LDs. CGI-58 is not required for this process. Thus, CGI-58 facilitates lipolysis in cooperation with perilipin and other factors, including lipases.

A case with full clinical manifestations of Dorfman-Chanarin syndrome

Abstract Dorfman-Chanarin syndrome (DCS), is a rare, autosomal recessive disorder associated with lipid metabolism. It is characterized by ichthyosiform nonbullous erythroderma, lipid vacuoles in peripheral leukocytes and variable involvement of organs. We report a Turkish man with the complete syndrome, who described family history of ichthyosis. To best of our knowledge this is the sixth case from Turkish origin to date. In addition to congenital ichthyosis he had also strabismus, horizantal nystagmus, bilateral neurosensory hearing loss, hepatomegaly and splenomegaly. Liver biopsy revealed hidrophic degeneration in hepatocytes, steatosis, enlargement and inflammation in portal areas and portal central fibrosis, consistent with cirrhosis. Write stained peripheral blood smear examination revealed lipid vacuoles in all of the neutrophils consistent with Jordan’s anomaly. We think that, it is essential to evaluate the peripheral blood smear of the patients with ichthyosis and also patients with DCS should be informed and warned about the results of consanquinous marriage.

An original mutation of CGI-58 in a new case of Dorfman–Chanarin syndrome

An original mutation of CGI-58 in a new case of Dorfman–Chanarin syndrome

Dorfman-Chanarin syndrome

A four-year-old girl was brought to the dermatology outpatient department with scaling all over the body since birth. She had history of episodic vomiting and abdominal distension. A dermatological diagnosis of lamellar ichthyosis was made. Abdominal examination revealed a nontender hepatomegaly, fatty liver on ultrasonography and deranged liver function tests. Peripheral blood smear showed lipid vacuoles in the granulocytes consistent with Jordans' anomaly. Similar lipid vacuoles were seen in the basal layer in skin biopsy. An inflammatory infiltrate, moderate fibrosis in the portal tract and diffuse severe fatty change in hepatocytes were seen in liver biopsy. The patient was diagnosed as a case of Dorfman-Chanarin syndrome.

The gene encoding adipose triglyceride lipase (PNPLA2) is mutated in neutral lipid storage disease with myopathy

Neutral lipid storage disease comprises a heterogeneous group of autosomal recessive disorders characterized by systemic accumulation of triglycerides in cytoplasmic droplets. Here we report a neutral lipid storage disease subgroup characterized by mild myopathy, absence of ichthyosis and mutations in both alleles of adipose triglyceride lipase (PNPLA2, also known as ATGL). Three of these mutations are predicted to lead to a truncated ATGL protein with an intact patatin domain containing the active site, but with defects in the hydrophobic domain. The block in triglyceride degradation was mimicked by short interfering RNA directed against ATGL. NLSDM is distinct from Chanarin-Dorfman syndrome, which is characterized by neutral lipid storage disease with ichthyosis, mild myopathy and hepatomegaly due to mutations in ABHD5 (also known as CGI-58).

Barrier Dysfunction and Pathogenesis of Neutral Lipid Storage Disease with Ichthyosis (Chanarin–Dorfman Syndrome)

Neutral lipid storage disease with ichthyosis (NLSDI; Chanarin-Dorfman syndrome) is an ichthyosiform syndrome, often associated with mutations in a lipid hydrolase, CGI-58. The presence of oil red O-positive, neutral lipid droplets in tissue biopsies, and/or in leukocytes on blood smears, coupled with a constellation of multisystem abnormalities and a pruritic ichthyosiform erythroderma, are together diagnostic of NLSDI. We investigated the pathogenesis of the ichthyosiform erythroderma in patients from three unrelated kindreds with a clinical diagnosis of NLSDI. Basal permeability barrier function and stratum corneum (SC) integrity were abnormal, but barrier recovery rates were faster than normal, as in atopic dermatitis. The basal barrier abnormality was linked to the secretion of lipid micro-inclusions, first segregated within lamellar bodies (LB), which then form a non-lamellar phase within the SC interstices, shown by combined ruthenium tetroxide post-fixation and lipid-retaining resin-white embedding. With colloidal lanthanum nitrate perfusion, excess water/solute movement was restricted to the SC interstices, and further localized to non-lamellar domains. Phase separation of excess stored lipid provides a unifying pathogenic mechanism not only for NLSDI, but also in several other inherited ichthyosiform disorders of lipid metabolism, such as recessive X-linked ichthyosis and type 2 Gaucher's disease.

Adipose triglyceride lipase-mediated lipolysis of cellular fat stores is activated by CGI-58 and defective in Chanarin-Dorfman Syndrome

Adipose triglyceride lipase (ATGL) was recently identified as an important triacylglycerol (TG) hydrolase promoting the catabolism of stored fat in adipose and nonadipose tissues. We now demonstrate that efficient ATGL enzyme activity requires activation by CGI-58. Mutations in the human CGI-58 gene are associated with Chanarin-Dorfman Syndrome (CDS), a rare genetic disease where TG accumulates excessively in multiple tissues. CGI-58 interacts with ATGL, stimulating its TG hydrolase activity up to 20-fold. Alleles of CGI-58 carrying point mutations associated with CDS fail to activate ATGL. Moreover, CGI-58/ATGL coexpression attenuates lipid accumulation in COS-7 cells. Antisense RNA-mediated reduction of CGI-58 expression in 3T3-L1 adipocytes inhibits TG mobilization. Finally, expression of functional CGI-58 in CDS fibroblasts restores lipolysis and reverses the abnormal TG accumulation typical for CDS. These data establish an important biochemical function for CGI-58 in the lipolytic degradation of fat, implicating this lipolysis activator in the pathogenesis of CDS.

Analysis of interaction partners for perilipin and ADRP on lipid droplets∗

Despite the critical roles of intracellular lipid droplets (LDs) in lipid storage and metabolism, little is known about the molecular mechanisms of their functions. Several protein components associated with the surface of LDs have been identified. A major one is perilipin in adipocytes and steroidogenic cells, whereas ADRP in most other cell types. They are loosely grouped as a small protein family sharing a common N-terminal motif, called the PAT domain. Perilipin regulates the breakdown of triacylglycerol in LDs via its phosphorylation. ADRP is characterized as a fatty acid binding protein and involved in lipid uptake and LD formation. For examining the functions of perilipin and ADRP at the molecular level, we performed yeast two-hybrid screening in this study, to find their functional partners. We identified CGI-58, a product of the causal gene of Chanarin-Dorfman syndrome (CDS), as an interactor for both perilipin and ADRP. Specific interaction between CGI-58 and perilipin was confirmed in a GST-pulldown assay and supported by fluorescence microscopic analyses. We further demonstrated that CGI-58 is principally located at the surface of LDs in 3T3-L1 cells, together with perilipin, and its expression is upregulated upon stimulation for adipocyte differentiation. Other than CGI-58, we also identified in yeast two-hybrid screening HSP86 and D52 tumor proteins as binding partners of perilipin, and IRG-47 of ADRP. These factors might be cooperated with perilipin and ADRP, and hence involved in membrane dynamics of LDs as well as the regulation of lipolysis on the surface of LDs.

Erythrokeratoderma variabilis-like ichthyosis in Chanarin-Dorfman syndrome

Neutral lipid storage disease (Chanarin-Dorfman syndrome) is an autosomal recessive metabolic disorder associated with congenital ichthyosis and a multisystemic accumulation of neutral lipids (lipid droplets) in various types of cells. The clinical presentation has been reported to correspond to that of nonbullous congenital ichthyosiform erythroderma. We report a 4-year-old boy presenting a generalized ichthyosiform disorder manifested by migrating scaly plaques alternating with areas of normal-looking skin, showing erythematous borders with sharp margins, clinically suggestive of erythrokeratoderma variabilis (EKV). A peripheral blood smear revealed cytoplasmic vacuoles in most granulocytes. Genetic studies from the patient and his parents revealed that the patient carried two different and novel mutations of the ABHD5 gene: a nonsense mutation in exon 6 (transmitted by the father) and an insertion/deletion in exon 4 (transmitted by the mother). Our observation demonstrates the clinical heterogeneity of the ichthyosiform dermatoses observed in Chanarin-Dorfman syndrome and widens the clinical range of conditions presenting migrating scaly plaques mimicking EKV.

Two New Mutations of the ABHD5 Gene in a New Adult Case of Chanarin Dorfman Syndrome: An Uncommon Lipid Storage Disease

Two New Mutations of the ABHD5 Gene in a New Adult Case of Chanarin Dorfman Syndrome: An Uncommon Lipid Storage Disease

Hepatobiliary and pancreatic: Dorfman–Chanarin syndrome

Hepatobiliary and pancreatic: Dorfman–Chanarin syndrome

CGI-58 Interacts with Perilipin and Is Localized to Lipid Droplets: POSSIBLE INVOLVEMENT OF CGI-58 MISLOCALIZATION IN CHANARIN-DORFMAN SYNDROME

Lipid droplets (LDs) are a class of ubiquitous cellular organelles that are involved in lipid storage and metabolism. Although the mechanisms of the biogenesis of LDs are still unclear, a set of proteins called the PAT domain family have been characterized as factors associating with LDs. Perilipin, a member of this family, is expressed exclusively in the adipose tissue and regulates the breakdown of triacylglycerol in LDs via its phosphorylation. In this study, we used a yeast two-hybrid system to examine the potential function of perilipin. We found direct interaction between perilipin and CGI-58, a deficiency of which correlated with the pathogenesis of Chanarin-Dorfman syndrome (CDS). Endogenous CGI-58 was distributed predominantly on the surface of LDs in differentiated 3T3-L1 cells, and its expression increased during adipocyte differentiation. Overexpressed CGI-58 tagged with GFP gathered at the surface of LDs and colocalized with perilipin. This interaction seems physiologically important because CGI-58 mutants carrying an amino acid substitution identical to that found in CDS lost the ability to be recruited to LDs. These mutations significantly weakened the binding of CGI-58 with perilipin, indicating that the loss of this interaction is involved in the etiology of CDS. Furthermore, we identified CGI-58 as a binding partner of ADRP, another PAT domain protein expressed ubiquitously, by yeast two-hybrid assay. GFP-CGI-58 expressed in non-differentiated 3T3-L1 or CHO-K1 cells was colocalized with ADRP, and the CGI-58 mutants were not recruited to LDs carrying ADRP, indicating that CGI-58 may also cooperate with ADRP.

Steatohepatitis and unsuspected micronodular cirrhosis in Dorfman-Chanarin syndrome with documented ABHD5 mutation

Mutation in ABHD5 causes Dorfman-Chanarin syndrome (DCS), a multisystem triglyceride storage disorder. Ultrastructural study of leukocytes confirmed DCS in a child homozygous for a novel ABHD5 mutation, with ichthyosis, developmental delay, and steatohepatitis with cirrhosis, manifest only as elevated aminotranferase levels. We recommend early assessment for liver disease in DCS.

Steatohepatitis and unsuspected micronodular cirrhosis in Dorfman-Chanarin syndrome with documented ABHD5 mutation

Steatohepatitis and unsuspected micronodular cirrhosis in Dorfman-Chanarin syndrome with documented ABHD5 mutation

Truncation of CGI-58 Protein Causes Malformation of Lamellar Granules Resulting in Ichthyosis in Dorfman-Chanarin Syndrome

Dorfman-Chanarin syndrome is a rare autosomal recessive inherited lipid storage disease characterized by ichthyosis, leukocyte lipid vacuoles, and involvement of several internal organs. Recently, CGI-58 mutations were identified as the cause of Dorfman-Chanarin syndrome. The physiologic roles of the CGI-58 protein and the pathomechanisms of Dorfman-Chanarin syndrome still remain to be clarified, however. The patient, a 16-y-old male, demonstrated ichthyosis, small ears, lipid vacuoles in his leukocytes, liver dysfunction, and mental retardation. Sequencing of CGI-58 revealed that the patient was homozygous for a novel nonsense mutation R184X, in exon 4. The putative truncated protein was 52.4% of the length of the normal CGI-58 polypeptide and lacked approximately 60% of the lipid binding region, 66.4% of the alpha/beta hydrolase folding segment of the polypeptide, and two of the CGI-58 catalytic triads, resulting in a significant loss of lipase/esterase/thioesterase activity. Electron microscopy revealed a large number of abnormal lamellar granules, a disturbed intercellular lamellar structure, and lipid vacuoles in the epidermis. These results suggested that CGI-58 protein is involved in the lipid metabolism of lamellar granules and that defective lipid production in lamellar granules caused by a CGI-58 protein deficiency is involved in the pathogenesis of ichthyosis in Dorfman-Chanarin syndrome.

Metabolic and drug-induced muscle disorders.

The inherited disorders of muscle metabolism affect both substrate utilization and the final intramitochondrial oxidation through the Krebs cycle and the respiratory chain. Almost every step of these complex biochemical pathways can be affected by inborn errors, whose expression depends on peculiar tissue-specific or systemic gene expression. This review updates current knowledge in this broad field. New inherited defects are still being discovered, such as the beta-enolase deficiency in glycogenosis type XIII and mutations in the gene encoding an esterase/lipase/thioesterase protein in Chanarin-Dorfman syndrome, a multisystem triglyceride storage disease. Therapeutic approaches to the metabolic myopathies are still lagging behind, although remarkable observations have been made on the rare coenzyme Q10 deficiency syndrome. However, transgenic animal models may offer the opportunity both to investigate muscle pathogenesis and explore therapeutic targets. Finally, human myotoxicity may provide novel paradigms for naturally occurring muscle disorders.

Mutations in CGI-58, the Gene Encoding a New Protein of the Esterase/Lipase/Thioesterase Subfamily, in Chanarin-Dorfman Syndrome

Chanarin-Dorfman syndrome (CDS) is a rare autosomal recessive form of nonbullous congenital ichthyosiform erythroderma (NCIE) that is characterized by the presence of intracellular lipid droplets in most tissues. We previously localized a gene for a subset of NCIE to chromosome 3 (designated "the NCIE2 locus"), in six families. Lipid droplets were found in five of these six families, suggesting a diagnosis of CDS. Four additional families selected on the basis of a confirmed diagnosis of CDS also showed linkage to the NCIE2 locus. Linkage-disequilibrium analysis of these families, all from the Mediterranean basin, allowed us to refine the NCIE2 locus to an approximately 1.3-Mb region. Candidate genes from the interval were screened, and eight distinct mutations in the recently identified CGI-58 gene were found in 13 patients from these nine families. The spectrum of gene variants included insertion, deletion, splice-site, and point mutations. The CGI-58 protein belongs to a large family of proteins characterized by an alpha/beta hydrolase fold. CGI-58 contains three sequence motifs that correspond to a catalytic triad found in the esterase/lipase/thioesterase subfamily. Interestingly, CGI-58 differs from other members of the esterase/lipase/thioesterase subfamily in that its putative catalytic triad contains an asparagine in place of the usual serine residue.

Dorfman-Chanarin syndrome (neutral lipid storage disease): new clinical features

Sir, Dorfman–Chanarin syndrome (DCS, neutral lipid storage disease, multisystem triglyceride storage disorder) is an autosomal recessive entity in which an excess of triacylglycerol accumulates in most cells, resulting from blockage in a recycling pathway that converts triglycerides to phospholipids.1 This syndrome is characterized by the association of congenital ichthyosis with the presence of lipid droplets within leucocytes (Jordans’ anomaly). Extracutaneous manifestations in DCS2 range between no extracutaneous involvement and multisystem involvement, usually without severe consequences for life. The recent observation that a diet poor in long‐chain fatty acids and enriched with medium‐chain fatty acids improved skin and liver manifestations in a boy suffering from this syndrome3 suggested that early initiation of such a diet might prevent systemic disturbances and improve skin alterations. A 37‐year‐old Spanish woman presented with congenital ichthyosis and muscular weakness. There was no consanguinity or familial cases of ichthyosis. Examination revealed large white plate‐like scales over her entire body surface, including flexural areas (Fig. 1a). No erythema was seen. The facial skin was tense, shiny and dry, with bilateral ectropion. There was total alopecia of the scalp. The palms and soles were hyperkeratotic, and transverse leuconychia was present (Fig. 1b). Skin biopsy showed ortho‐ and para‐keratosis and a mild dermal perivascular inflammatory infiltrate. Oil‐red‐O staining demonstrated vacuoles of neutral lipids in the stratum corneum, stratum granulosum and sweat ducts. She had short stature (height 1·47 m). Liver and spleen were not enlarged. Neurological examination and psychomotor development were normal, and there were no hearing abnormalities. Ophthalmological examination revealed a total cataract of the left eye and bilateral corneal leucomas. Visual acuity was diminished in her right eye (0·4), with myopia and astigmatism. Blood count, urea, creatinine, electrolytes, glucose, hepatic enzymes, cholesterol, triglycerides, creatine phosphokinase, aldolase, antinuclear antibodies and urinalysis were normal. Urinary creatine was elevated (181 mg in 24 h). A peripheral blood smear showed two to five lipid vacuoles in most granulocytes (Fig. 1c). Abdominal ultrasound, thoracic scan and abdominal scan were normal. Consent for muscle biopsy and electromyography was not given. A diet poor in long‐chain fatty acids and enriched with medium‐chain fatty acids, and acitretin 30 mg daily, were prescribed, with improvement of skin and muscle manifestations.

A cause of fatty liver: neutral lipid storage disease with ichthyosis--electron microscopic findings.

A cause of fatty liver: neutral lipid storage disease with ichthyosis--electron microscopic findings.

Neutral lipid storage disease: a genetic disorder with abnormalities in the regulation of phospholipid metabolism

Neutral lipid storage disease (NLSD) is an autosomal recessive disorder characterized by the presence of numerous lipid droplets in virtually all tissues examined. The increased cellular triacylglycerol content results from defective recycling of triacylglycerol-derived diacylglycerol to phospholipids (Igal, R. A. and R. A. Coleman. 1996. J. Biol. Chem.271: 16644–16651). In order to determine whether de novo glycerolipid synthesis is also altered in NLSD, we compared the ability of normal human skin fibroblasts and fibroblasts from a patient with NLSD to incorporate phospholipid precursors into cell lipids. NLSD cells had increased rates of incorporation of [14C]oleic acid and [3H]glycerol into triacylglycerol and all phospholipid species except phosphatidylethanolamine. However, the cell content of each phospholipid species was similar in control and NLSD cells, indicating a higher turnover rate in NLSD cells for phosphatidylcholine, phosphatidylinositol, phosphatidylserine, and sphingomyelin. Labeling with [14C]choline and [14C]ethanolamine confirmed the increase in the rate of phosphatidylcholine synthesis and the decreased rate of phosphatidylethanolamine synthesis through their respective CDP pathways. The activities of the major regulatory enzymes of triacylglycerol, phosphatidylcholine, and phosphatidylethanolamine biosynthesis were similar in control and NLSD cells. Taken as a whole, this study provides strong evidence for an underlying regulatory defect in NLSD that alters the rates of synthesis and degradation of the major cellular phospholipids.—Igal, R. A., and R. A. Coleman. Neutral lipid storage disease: a genetic disorder with abnormalities in the regulation of phospholipid metabolism. J. Lipid Res. 1998. 39: 31–43.