LCAT Deficiency Research Articles

Role of LPX in the development of renal disease in LCAT deficiency

Role of LPX in the development of renal disease in LCAT deficiency

9. AAV-Mediated Gene Transfer as a Therapeutic Approach for Familial LCAT Deficiency

Lecithin Cholesterol Acyltransferase (LCAT) is a plasma enzyme secreted by the liver and responsible for esterification of free cholesterol to cholesteryl ester in the blood compartment. Familial LCAT deficiency (FLD) is an autosomal recessive disorder caused by loss-of-function mutations of the LCAT gene and is characterized by extremely low levels of high density lipoprotein cholesterol (HDL-C), anemia, corneal opacities, and chronic progressive renal disease leading to end-stage kidney disease as early as the third decade of life. Currently, there is no specific treatment for FLD. The fact that patients with even very low levels of plasma LCAT activity do not develop kidney disease make FLD an attractive candidate for liver-directed gene therapy with adeno-associated viral (AAV) vectors. LCAT is activated by apolipoprotein A-I (apoA-I) and this interaction is highly species specific. Therefore, LCAT KO mice were crossed with human apoA-I transgenic (hApoAITg) mice in order to facilitate experiments involving expression of human LCAT. LCAT KO/hApoAI transgenic mice received a single IP injection of AAV8 encoding human LCAT in a dose range of 5e10 GC/kg to 5e11 GC/kg. HDL-C increased in a dose-dependent fashion, from 32 mg/dL in the control group to 103 mg/dL at the dose of 1.5e11 GC/kg and 473 mg/dL at the dose of 5e11 GC/kg. Plasma% cholesterol ester, a sensitive indicator of LCAT activity, was normalized at the low dose of 1.5e11 GC/kg. Thus, we demonstrate that low doses of AAV8-hLCAT provide stable expression of the LCAT enzyme and correct dyslipidemia in a ‘humanized’ mouse model of LCAT deficiency. Studies designed to demonstrate prevention of kidney disease in this mouse model are ongoing. These proof-of-concept studies provide a foundation for further development of AAV-based gene therapy for familial LCAT deficiency.

Distinct Roles of Apolipoprotein A1 and Lecithin‐Cholesterol Acyltransferase in Lipopolysaccharide‐Induced Inflammation

AimWe studied lipopolysaccharide (LPS)‐induced inflammation under conditions of Apoa1‐ or Lcat‐ deficiency.MethodsRAW 264.7 macrophages, whole blood and peritoneal macrophages from C57BL/6 (WT), Lcat‐deficient (Lcat‐/‐) and Apoa1‐deficient (Apoa1‐/‐) mice were challenged with LPS and plasma TNFα was measured. WT, Lcat‐/‐ and Apoa1‐/‐ mice were challenged intravenously with LPS and plasma cytokines were monitored. WT, Lcat‐/‐ and Apoa1‐/‐ leukocytes were phenotyped using FACS.ResultsSerum and HDL from Lcat‐/‐ and Apoa1‐/‐ mice display reduced capacity to attenuate the LPS‐induced TNFα response, in vitro. Blood from Lcat‐/‐ and Apoa1‐/‐ mice show an increased TNFα response when stimulated with LPS ex vivo, with more pronounced effect for Lcat‐/‐ blood. Lcat‐/‐ and Apoa1‐/‐ mice show a markedly enhanced and prolonged LPS‐induced inflammatory response in vivo. LCAT‐deficiency is associated with leukocytosis, increased CD11b+/Ly‐6Cmed monocyte and CD4+ lymphocyte levels, but low CD11b+/Ly‐6Chi monocyte levels and CD11b expression. Similarly, peritoneal macrophages from Lcat‐/‐ mice show reduced LPS‐induced TΝFα response. In contrast, APOA1 deficiency is associated with decreased leukocyte, CD11b+/Ly‐6Clo/med/hi monocyte and CD4+, CD8+ lymphocyte levels.ConclusionsLCAT deficiency renders leukocytes less responsive to LPS, but increases their number in circulation, while APOA1 deficiency results in fewer but more responsive leukocytes.

Structure and function of lysosomal phospholipase A2 and lecithin:cholesterol acyltransferase.

Lysosomal phospholipase A2 (LPLA2) and lecithin:cholesterol acyltransferase (LCAT) belong to a structurally uncharacterized family of key lipid metabolizing enzymes responsible for lung surfactant catabolism and for reverse cholesterol transport, respectively. Whereas LPLA2 is predicted to underlie the development of drug-induced phospholipidosis, somatic mutations in LCAT cause fish eye disease and familial LCAT deficiency. Here we describe several high resolution crystal structures of human LPLA2 and a low resolution structure of LCAT that confirms its close structural relationship to LPLA2. Insertions in the α/β hydrolase core of LPLA2 form domains that are responsible for membrane interaction and binding the acyl chains and head groups of phospholipid substrates. The LCAT structure suggests the molecular basis underlying human disease for most of the known LCAT missense mutations, and paves the way for rational development of new therapeutics to treat LCAT deficiency, atherosclerosis and acute coronary syndrome.

A robust all-atom model for LCAT generated by homology modeling

LCAT is activated by apoA-I to form cholesteryl ester. We combined two structures, phospholipase A2 (PLA2) that hydrolyzes the ester bond at the sn-2 position of oxidized (short) acyl chains of phospholipid, and bacteriophage tubulin PhuZ, as C- and N-terminal templates, respectively, to create a novel homology model for human LCAT. The juxtaposition of multiple structural motifs matching experimental data is compelling evidence for the general correctness of many features of the model: i) The N-terminal 10 residues of the model, required for LCAT activity, extend the hydrophobic binding trough for the sn-2 chain 15-20 Å relative to PLA2. ii) The topography of the trough places the ester bond of the sn-2 chain less than 5 Å from the hydroxyl of the catalytic nucleophile, S181. iii) A β-hairpin resembling a lipase lid separates S181 from solvent. iv) S181 interacts with three functionally critical residues: E149, that regulates sn-2 chain specificity, and K128 and R147, whose mutations cause LCAT deficiency. Because the model provides a novel explanation for the complicated thermodynamic problem of the transfer of hydrophobic substrates from HDL to the catalytic triad of LCAT, it is an important step toward understanding the antiatherogenic role of HDL in reverse cholesterol transport.

HDL genetic defects.

High density lipoprotein cholesterol (HDL-C) and its related apolipoproteins form part of the reverse cholesterol transport system that removes excessive cholesterol from the periphery to the liver. Many transport proteins and enzymes that are involved in this process are susceptible to genetic defects that influence plasma HDL-C concentrations and HDL function. The HDL-C concentration in the blood may not be as important as the function of this lipid fraction. The genetic defects affecting plasma HDL-C concentrations do not always show a consistent relationship with atherosclerosis. Familial hypoalphalipoproteinaemia is associated with mutations in genes responsible for the transport proteins or the enzymes involved in the biogenesis of HDL-C. Inheritance of a Milano mutation of apolipoprotein A1 decreases the risk of atherosclerotic disease despite low circulating levels of HDL-C. Tangier disease and Fish Eye disease are caused by mutations in the ATP binding cassette A1 (ABCA1), a transport protein, and lecithin cholesterol acyl transferase (LCAT), an enzyme, involved in the esterification of cholesterol, respectively. Patients with these conditions have very low levels of HDL-C concentration. The association between both these conditions and the risk of cardiovascular disease (CVD) is variable and inconsistent. Understanding the molecular mechanism of HDL biogenesis not only helped in defining the pathophysiology of low and high HDL-C syndromes, but also in developing new treatment options to raise HDL-C levels.

Corneal dystrophies related to cholesterol metabolism (fish eye and Schnyder dystrophy)

AbstractSchnyder corneal dystrophy (SCD) is a rare corneal dystrophy characterized by abnormally increased deposition of cholesterol and phospholipids in the cornea. SCD is inherited as an autosomal dominant trait with high penetrance and has been mapped to the UBIAD1 gene on chromosome 1p36.3. Diseases of systemic lipid metabolism that cause corneal opacification, such as lecithin‐ cholesterol acyltransferase (LCAT) deficiency and fish eye disease (FED), should be considered in the differential diagnosis. LCAT deficiency and FED are both entities that result from deficiency of the LCAT enzyme. LCAT deficiency is defined by deficient LCAT activity towards HDL and LDL whereas FED is defined by decreased LCAT activity against HDL only. When comparing LCAT deficiency and FED to SCD, there are numerous differences to help distinguish all 3 entities. Both LCAT deficiency and FED are inherited in an autosomal recessive manner while SCD has autosomal dominant inheritance. Systemically, SCD has a normal level of LCAT enzyme activity and normal HDL levels. Both LCAT deficiency and FED show abnormally low levels of HDL. SCD is often associated with a true arcus lipoides, unlike FED and LCAT deficiency.

LCAT deficiency does not impair amyloid metabolism in APP/PS1 mice

A key step in plasma HDL maturation from discoidal to spherical particles is the esterification of cholesterol to cholesteryl ester, which is catalyzed by LCAT. HDL-like lipoproteins in cerebrospinal fluid (CSF) are also spherical, whereas nascent lipoprotein particles secreted from astrocytes are discoidal, suggesting that LCAT may play a similar role in the CNS. In plasma, apoA-I is the main LCAT activator, while in the CNS, it is believed to be apoE. apoE is directly involved in the pathological progression of Alzheimer's disease, including facilitating β-amyloid (Aβ) clearance from the brain, a function that requires its lipidation by ABCA1. However, whether apoE particle maturation by LCAT is also required for Aβ clearance is unknown. Here we characterized the impact of LCAT deficiency on CNS lipoprotein metabolism and amyloid pathology. Deletion of LCAT from APP/PS1 mice resulted in a pronounced decrease of apoA-I in plasma that was paralleled by decreased apoA-I levels in CSF and brain tissue, whereas apoE levels were unaffected. Furthermore, LCAT deficiency did not increase Aβ or amyloid in APP/PS1 LCAT(-/-) mice. Finally, LCAT expression and plasma activity were unaffected by age or the onset of Alzheimer's-like pathology in APP/PS1 mice. Taken together, these results suggest that apoE-containing discoidal HDLs do not require LCAT-dependent maturation to mediate efficient Aβ clearance.

Acquired lecithin:cholesterol acyltransferase deficiency as a major factor in lowering plasma HDL levels in chronic kidney disease.

It has been suggested that a low plasma high-density lipoprotein cholesterol (HDL-C) level contributes to the high cardiovascular disease risk of patients with chronic kidney disease (CKD), especially those undergoing haemodialysis (HD). The present study was conducted to gain further understanding of the mechanism(s) responsible for the low HDL-C levels in patients with CKD and to separate the impact of HD from that of the underlying CKD. Plasma lipids and lipoproteins, HDL subclasses and various cholesterol esterification parameters were measured in a total of 248 patients with CKD, 198 of whom were undergoing HD treatment and 40 healthy subjects. Chronic kidney disease was found to be associated with highly significant reductions in plasma HDL-C, unesterified cholesterol, apolipoprotein (apo)A-I, apoA-II and LpA-I:A-II levels in both CKD cohorts (with and without HD treatment). The cholesterol esterification process was markedly impaired, as indicated by reductions in plasma lecithin:cholesterol acyltransferase (LCAT) concentration and activity and cholesterol esterification rate, and by an increase in the plasma preβ-HDL content. HD treatment was associated with a further lowering of HDL levels and impaired plasma cholesterol esterification. The plasma HDL-C level was highly significantly correlated with LCAT concentration (R=0.438, P<0.001), LCAT activity (R=0.243, P<0.001) and cholesterol esterification rate (R=0.149, P=0.031). Highly significant correlations were also found between plasma LCAT concentration and levels of apoA-I (R=0.432, P<0.001), apoA-II (R=0.275, P<0.001), LpA-I (R= 0.326, P<0.001) and LpA-I:A-II (R=0.346, P< 0.001). Acquired LCAT deficiency is a major cause of low plasma HDL levels in patients with CKD, thus LCAT is an attractive target for therapeutic intervention to reverse dyslipidaemia, and possibly lower the cardiovascular disease risk in these patients.

High-density lipoprotein metabolism, composition, function, and deficiency.

To examine the recent advances in our knowledge of HDL metabolism, composition, function, and coronary heart disease (CHD), as well as marked HDL deficiency states because of mutations in the apolipoprotein (apo) A-I, ATP-binding cassette transfer protein A1 and lecithin cholesterol acyltransferase (LCAT) gene loci. It has been documented that apoA-I, myeloperoxidase and paraoxonase 1 (PON1) form a complex in HDL that is critical for HDL binding and function. Myeloperoxidase has a negative impact on HDL function, whereas PON1 has a beneficial effect. Patients who lack apoA-I develop markedly premature CHD. Patients who lack ATP-binding cassette transfer protein A1 transporter function have only very small discoidal preβ-1 HDL, and develop hepatosplenomegaly, intermittent neuropathy and premature CHD, although significant heterogeneity for these disorders has been reported. Patients with LCAT deficiency have abnormal small discoidal LDLs and HDL particles, and develop kidney failure. Enzyme replacement therapy is being developed for the latter disorder. Recent data indicates that proteins other than apoA-I and apoA-II such as MPO and PON1 have important effects on HDL function. There has been considerable recent progress made in our understanding of HDL protein content and function.

A Kindred with Fish Eye Disease, Corneal Opacities, Marked High Density Lipoprotein Defieciency, and Statin Therapy†

A Kindred with Fish Eye Disease, Corneal Opacities, Marked High Density Lipoprotein Defieciency, and Statin Therapy†

A kindred with fish eye disease, corneal opacities, marked high-density lipoprotein deficiency, and statin therapy

A kindred affected with fish eye disease (FED) from Oklahoma is reported. Two probands with corneal opacification had mean levels of high-density lipoprotein (HDL) cholesterol (C), apolipoprotein (apo) A-I, and apoA-I in very large alpha-1 HDL particles that were 9%, 17%, and 5% of normal, whereas their parents and 1 sibling had values that were 61%, 77%, and 72% of normal. The probands had no detectable lipoprotein-X, and had mean low-density lipoprotein cholesterol (LDL-C) and triglyceride levels that were elevated. Their mean lecithin cholesterol acyltransferase (LCAT) activities, cholesterol esterification rates, and free cholesterol levels were 8%, 42%, and 258% of normal, whereas their parents and 1 sibling had values that were 55%, 49%, and 114% of normal. The defect was due to 1 common variant in the LCAT gene in exon 1: c101t causing a proline34leucine substitution and a novel mutation c1177t causing a threonine37methionine substitution, with the former variant being found in the father and 1 sibling, and the latter mutation being found in the mother, and both mutations being present in the 2 probands. FED is distinguished from familial LCAT deficiency (FLD) by the lack of anemia, splenomegaly, and renal insufficiency as well as normal or increased LDL-C. Both FLD and FED cases have marked HDL deficiency and corneal opacification, and FED cases may have premature coronary heart disease in contrast to FLD cases. Therapy, using presently available agents, in FED should be to optimize LDL-C levels, and 1 proband responded well to statin therapy. The investigational use of human recombinant LCAT as an enzyme source is ongoing.

Abstract 18673: Human Enzyme Replacement Therapy in a Patient With Familial Lecithin Cholesterol Acyltransferase Deficiency: Rapid Appearance of Normal Appearing HDL

Introduction: Lecithin cholesterol acyltransferase (LCAT) is a liver-derived enzyme in plasma that catalyzes the formation of cholesteryl esters (CE). Mutations in the human LCAT gene result in familial LCAT deficiency (FLD), characterized by absent high-density lipoprotein cholesterol (HDL-C), corneal opacities, anemia, and severe renal disease. Replacing the defective enzyme should restore the normal level of plasma CE (LCAT product) and lead to the appearance of alpha HDL (α-HDL) lipoproteins. Aim: To assess the formation of CE and the appearance of HDL over time in a FLD patient with recombinant human LCAT (ACP-501). Methods: A FLD subject with very low HDL-C (&lt;5 mg/dL), corneal opacities, stage 4/5 renal disease, anemia, and splenomegaly received a 1 hour infusion of recombinant human LCAT (rhLCAT) and serial plasma samples were collected over 7 days. Results: HDL-C increased from &lt;5 mg/dL to 17 mg/dL at 8 hours and remained measurable for 7 days. CE increased rapidly, doubling in concentration within 2 hours, and sustained a higher concentration than HDL over 7 days. An increase in LDL-C was delayed about 6 hours suggesting CETP transfer of CE from HDL to LDL. Small sized particles were rapidly converted to HDL2 and HDL3 sized particles on FPLC. Large α-HDL appeared on agarose electrophoresis within 30 minutes. ApoAI Western blot analysis confirmed the rapid generation of apoAI staining α-HDL. α-1, 2 and 3 particles rapidly appeared by 2-D gel electrophoresis. Filipin stained 1-D gels revealed large α particles that persisted for 7 days. α-4 sized particles decreased as α-HDL formed. Sudan stained 1-D gels showed the generation of CE-rich α-HDL. Conclusions: The infusion of rhLCAT (ACP-501) in a lecithin cholesterol acyltransferase deficient subject with no HDL rapidly generated CE and α-HDL with evidence of sustained activity over 7 days. CE appeared in LDL suggesting the transfer of CE from HDL to LDL by CETP. The lipid and lipoprotein changes are consistent with normal HDL maturation.

Characteristic kidney pathology, gene abnormality and treatments in LCAT deficiency

Lecithin:cholesterol acyltransferase (LCAT) is a key enzyme involved in reverse cholesterol transport from the peripheral tissues to the liver. LCAT deficiency, in which this enzyme is congenitally absent, is a genetic disease that impairs the esterification of free cholesterol in the plasma, leading to accumulation of phospholipids, including lecithin, in the organs of the body; the clinical manifestations include corneal opacities, normochromic anemia, renal disorder, etc. The prognosis is determined by the degree of renal dysfunction, and renal biopsy specimens reveal characteristic light- and electron-microscopic findings. The disease, transmitted by autosomal recessive inheritance, is extremely rare. There have only been 88 gene mutations of the LCAT gene reported around the world, and 13 of them are from Japan. One of the characteristics of LCAT deficiency is the strong correlations among the patterns, extent and phenotypes of these gene mutations.

Recombinant human LCAT normalizes plasma lipoprotein profile in LCAT deficiency

Lecithin:cholesterol acyltransferase (LCAT) is the enzyme responsible for cholesterol esterification in plasma. Mutations in the LCAT gene leads to two rare disorders, familial LCAT deficiency and fish-eye disease, both characterized by severe hypoalphalipoproteinemia associated with several lipoprotein abnormalities. No specific treatment is presently available for genetic LCAT deficiency. In the present study, recombinant human LCAT was expressed and tested for its ability to correct the lipoprotein profile in LCAT deficient plasma. The results show that rhLCAT efficiently reduces the amount of unesterified cholesterol (−30%) and promotes the production of plasma cholesteryl esters (+210%) in LCAT deficient plasma. rhLCAT induces a marked increase in HDL-C levels (+89%) and induces the maturation of small preβ-HDL into alpha-migrating particles. Moreover, the abnormal phospholipid-rich particles migrating in the LDL region were converted in normally sized LDL.

Abstract 450: Effects of Multiple Infusions of Recombinant LCAT on Plasma and Tissue Lipids and Expression of Genes Involved in Atherogenesis

BACKGROUND LCAT, an enzyme that converts small, discoidal HDL to spherical HDL particles by esterifying free cholesterol, plays a key role in reverse cholesterol transport (RCT). LCAT replacement therapy holds promise for treatment of familial LCAT deficiency, as well as a possible acute treatment in patients with Acute Coronary Syndrome. HYPOTHESIS Injecting recombinant LCAT into LCAT-deficient mice expressing human apoA-I will increase plasma HDL-C levels, improve reverse cholesterol transport, and alter expression of genes involved in atherosclerosis. METHODS We injected recombinant human LCAT or vehicle intravenously daily for 4 days into LCAT-KO x human apoA-I transgenic mice (n=4-6 in each group). The dose of rLCAT was approximately 10-fold larger in terms of mass than the endogenous levels of mouse LCAT. Lipids in plasma and tissues, as well as expression of 84 cholesterol metabolism genes in control and experimental groups were measured. RESULTS Total cholesterol (TC) in plasma increased from 80 (vehicle) to 473 mg/dl (LCAT) ( P &lt;0.002) after rLCAT treatment. Cholesteryl esters (CE) increased from 8 (vehicle) to 339 mg/dl (LCAT). Lipids were localized mostly in the HDL fraction. The accumulation of lipids in plasma was accompanied by decreased lipid content in peripheral tissues.TC in aorta, heart, kidney and spleen decreased, by 15%; 27%; 13% and 27%, respectively ( P &lt;0.05). In kidney, lung and spleen, CE decreased from 17; 18 and 28 nmol/mg dry weight, respectively, to being below detection. LCAT injections did not significantly alter liver lipid content, but did significantly increase expression of cholesterol synthesis genes (Dhcr7, Hmgcr, Mdv, Mvk and Tm7sf2) and decreased expression of the cholesterol efflux gene Abcg1.To further investigate the potential of interaction between apoA-I and LCAT, we also studied apoA-I-Tg x LCAT-Tg double transgenic mice. Their TC in plasma reached more than 1300 mg/mL, with similar gene expression changes observed in hapoA-I transgenic mice but more pronounced. CONCLUSIONS The results show the feasibility of using recombinant LCAT as a therapy for LCAT deficiency and for other diseases associated with accumulation of cholesterol in tissues.

Nephrotic Syndrome Caused by Immune-Mediated Acquired LCAT Deficiency

Lecithin-cholesterol acyltransferase (LCAT) is an enzyme involved in maintaining cholesterol homeostasis. In familial LCAT deficiency (FLD), abnormal lipid deposition causes renal injury and nephrotic syndrome, frequently progressing to ESRD. Here, we describe a 63-year-old Japanese woman with no family history of renal disease who presented with nephrotic syndrome. The laboratory data revealed an extremely low level of serum HDL and undetectable serum LCAT activity. Renal biopsy showed glomerular lipid deposition with prominent accumulation of foam cells, similar to the histologic findings of FLD. In addition, she had subepithelial electron-dense deposits compatible with membranous nephropathy, which are not typical of FLD. A mixing test and coimmunoprecipitation study demonstrated the presence of an inhibitory anti-LCAT antibody in the patient's serum. Immunohistochemistry and immunofluorescence detected LCAT along parts of the glomerular capillary walls, suggesting that LCAT was an antigen responsible for the membranous nephropathy. Treatment with steroids resulted in complete remission of the nephrotic syndrome, normalization of serum LCAT activity and HDL level, and disappearance of foam cell accumulation in renal tissue. In summary, inhibitory anti-LCAT antibody can lead to glomerular lesions similar to those observed in FLD.

Amelioration of circulating lipoprotein profile and proteinuria in a patient with LCAT deficiency due to a novel mutation (Cys74Tyr) in the lid region of LCAT under a fat-restricted diet and ARB treatment

Familial lecithin-cholesterol acyltransferase (LCAT) deficiency is a hereditary disease characterized by an abnormal lipid profile, corneal opacity, anemia and progressive renal disease. We report a patient with complete loss of LCAT activity due to a novel lcat gene mutation of Cys74Tyr in the lid region of LCAT protein. Esterification of cholesterol in this patient was disturbed by disruption of a substrate binding loop of Cys50-Cys74 in LCAT protein. She had progressive renal dysfunction, proteinuria, corneal opacity, anemia and an abnormal lipid profile. Her serum lipids showed a significant increase in abnormal lipoproteins at the original position in agarose gel electrophoresis and VLDL-cholesterol, and a severe decrease in serum HDL-cholesterol. Lipoprotein analyzes also revealed the presence of an abnormal midband lipoprotein, and a maturation disturbance of HDL particles. Renal function and proteinuria improved following the adoption of a fat-restricted diet and administration of an angiotensin II receptor blocker. The abnormal lipoproteins also decreased after this treatment.

Lipoprotein X: clinical implications

In this issue of the journal, Stepien et al. describe a case of serum lipoprotein X (Lp-X) in a patient with cholestasis and severe hyperlipidaemia. This abnormal lipoprotein is rich in phospholipid and unesterified cholesterol. The main protein component is albumin within the core and apolipoprotein C upon the surface. Unlike low-density lipoprotein (LDL), Lp-X does not contain apolipoprotein B nor is it removed by the LDL receptor, but Lp-X is cleared by the reticuloendothelial system and the kidneys. Lp-X is about 50 nm in diameter and its major component bile acid is lithocholic. Early studies reported the presence of Lp-X in the serum of patients with cholestasis. The failure of Lp-X to exert feedback inhibition on cholesterol synthesis may contribute to the mechanism of hypercholesterolaemia in obstructive jaundice. However, the presence of serum Lp-X does not facilitate the distinction between intrahepatic and extrahepatic cholestasis. Bile lipoprotein is a precursor of Lp-X and in cholestasis this refluxes into the plasma pool and binds to albumin to form Lp-X. Serum Lp-X usually disappears soon after the cholestasis is relieved. The ratio of plasma bile salts to albumin is important for the integrity of Lp-X and it is catabolized by the action of phospholipase which helps convert lecithin into lysolecithin. In graft versus host disease involving the liver and with cholestasis plasma Lp-X can result in severe hypercholesterolaemia, xanthelasma and cholesteroloma. Lp-X can also be found in cholestasis of pregnancy and primary biliary cirrhosis (PBC). The case presented by Stepien et al. confirms these earlier observations of raised serum Lp-X in cholestasis. Increased serum cholesterol concentrations occur in PBC and this is largely due to the presence of Lp-X, although it is debatable whether this confers increased cardiovascular risk. Indeed, Lp-X may not be atherogenic and data suggest it may have antioxidant LDL activity. Additionally, Lp-X is not significantly taken up by macrophages but is taken up by hepatocytes. Sometimes severe and prolonged cholestasis can result in very high serum Lp-X concentrations resulting in severe hypercholesterolaemia and tendon xanthomata. Complete regression of these xanthomata has been described by treatment with apheresis, which the authors stated should be considered possible first-choice treatment for patients with massive Lp-X accumulation due to cholestasis. Statin therapy may be ineffective in lowering Lp-X. Lecithin-cholesterol acyltransferase (LCAT) is an enzyme responsible for cholesterol esterification in plasma. Lp-X is found in LCAT deficiency which is characterized by low concentrations of high-density lipoprotein (HDL) as well as anaemia, corneal opacities, proteinuria and chronic kidney disease. The Lp-X found in LCAT deficiency may contribute to the renal disease observed in this condition by stimulating monocyte renal infiltration. Proteoglycans have an important role in regulating the uptake of Lp-X into kidney mesangial cells and this mechanism is independent of the scavenger receptor. Guerin et al. reported that in LCAT deficiency the presence of Lp-X is correlated with a progressive change in renal function. In addition to cholestasis and LCAT deficiency, Lp-X may also be found in the plasma after intravenous infusion of Intralipid emulsion such as occurs with total parenteral nutrition. Intralipid administration can elicit an increase in serum total cholesterol (mainly unesterified), phospholipid and LDL. The Lp-X associated with Intralipid infusion seems to occur when phospholipid clearance is exceeded, resulting in phospholipid accumulation with unesterified cholesterol. Pseudohyponatraemia has also been described in some patients with severe hypercholesterolaemia secondary to raised Lp-X. Thus the presence of Lp-X should be considered in pseudohyponatraemia associated with cholestasis. Sivakumar et al. described not only pseudohyponatraemia, but also pseudohypokalaemia and pseudohypochloridaemia in the presence of high concentrations of Lp-X. Various researchers have also found assay bias for serum LDL-cholesterol in the presence of Lp-X. This is an important practical point when it comes to interpreting LDL cholesterol results in severely cholestatic serum samples. Lp-X possesses strong aggregatory properties and can bind to alkaline phosphatase (ALP) producing a Lp-X-ALP complex. This complex may be found in cholestasis; typically that evoked by hepatic malignancy. The Lp-X-ALP complex is also associated with the presence of high-molecular mass ALP and the liver isoenzyme of ALP. Brocklehurst et al. described a stationary band of ALP using starch gel electrophoresis of serum samples from patients with biliary obstruction which was thought to be due to ALP binding to Lp-X. The strong aggregating ability of Lp-X may be important as it can bind and remove free cholesterol from the circulation. There are various assays available for measuring serum Lp-X. Agarose gel electrophoresis utilizes the cathodal

LCAT deficiency in mice is associated with a diminished adrenal glucocorticoid function

In vitro studies have suggested that HDL and apoB-containing lipoproteins can provide cholesterol for synthesis of glucocorticoids. Here we assessed adrenal glucocorticoid function in LCAT knockout (KO) mice to determine the specific contribution of HDL-cholesteryl esters to adrenal glucocorticoid output in vivo. LCAT KO mice exhibit an 8-fold higher plasma free cholesterol-to-cholesteryl ester ratio (P < 0.001) and complete HDL-cholesteryl ester deficiency. ApoB-containing lipoprotein and associated triglyceride levels are increased in LCAT KO mice as compared with C57BL/6 control mice (44%; P < 0.05). Glucocorticoid-producing adrenocortical cells within the zona fasciculata in LCAT KO mice are devoid of neutral lipids. However, adrenal weights and basal corticosterone levels are not significantly changed in LCAT KO mice. In contrast, adrenals of LCAT KO mice show compensatory up-regulation of genes involved in cholesterol synthesis (HMG-CoA reductase; 516%; P < 0.001) and acquisition (LDL receptor; 385%; P < 0.001) and a marked 40-50% lower glucocorticoid response to adrenocorticotropic hormone exposure, endotoxemia, or fasting (P < 0.001 for all). In conclusion, our studies show that HDL-cholesteryl ester deficiency in LCAT KO mice is associated with a 40-50% lower adrenal glucocorticoid output. These findings further highlight the important novel role for HDL as cholesterol donor for the synthesis of glucocorticoids by the adrenals.