A novel pathogenic variant in LCAT causing FLD. A case report.

BACKGROUND: Fish-eye disease (FED) is due to a partial deficiency in LCAT activity. Nevertheless, Familial lecithin-cholesterol acyltransferase deficiency (FLD), also called Norum disease, appears when the deficiency is complete. They are both rare genetic disorders inherited in an autosomal recessive manner. Clinical signs include decreased circulating HDL cholesterol and dense corneal opacity. Kidney injuries also affect patients suffering from FLD. The diagnosis of FLD is based on the presence of characteristic signs and symptoms and confirmed by genetic testing. CASE PRESENTATION: We present a case of a 63-year-old man showing an altered lipid profile with low HDL cholesterol, chronic kidney disease (CKD) and corneal disorders. He was referred to genetic counseling in order to discard genetic LCAT deficiency due to decreased visual acuity caused by corneal opacity. A massive DNA sequencing was conducted using a multigene panel associated with lipid metabolism disturbances. RESULTS AND GENETIC FINDINGS: Two likely pathogenic variants in LCAT were identified and later confirmed by Sanger sequencing. Both (c.491 G > A and c.496 G > A) were missense variants that originated an amino acid substitution (164Arginine for Histidine and 166Alanine for Threonine, respectively) modifying the protein sequence and its 3D structure. CONCLUSIONS: FLD and FED sharing common biochemical features, and the existence of other diseases with similar clinical profiles underline the need for a timely differential diagnosis aiming to address patients to preventive programs and future available therapies. This case, added to the reduced number of publications previously reported regarding FLD and FED, contributes to better understanding the genetic characteristics, clinical features, and diagnosis of these syndromes.

Physaria fendleri and Ricinus communis lecithin:cholesterol acyltransferase-like phospholipases selectively cleave hydroxy acyl chains from phosphatidylcholine.

Production of hydroxy fatty acids (HFAs) in transgenic crops represents a promising strategy to meet our demands for specialized plant oils with industrial applications. The expression of Ricinus communis (castor) OLEATE 12-HYDROXYLASE (RcFAH12) in Arabidopsis has resulted in only limited accumulation of HFAs in seeds, which probably results from inefficient transfer of HFAs from their site of synthesis (phosphatidylcholine; PC) to triacylglycerol (TAG), especially at the sn-1/3 positions of TAG. Phospholipase As (PLAs) may be directly involved in the liberation of HFAs from PC, but the functions of their over-expression in HFA accumulation and distribution at TAG in transgenic plants have not been well studied. In this work, the functions of lecithin:cholesterol acyltransferase-like PLAs (LCAT-PLAs) in HFA biosynthesis were characterized. The LCAT-PLAs were shown to exhibit homology to LCAT and mammalian lysosomal PLA2 , and to contain a conserved and functional Ser/His/Asp catalytic triad. In vitro assays revealed that LCAT-PLAs from the HFA-accumulating plant species Physaria fendleri (PfLCAT-PLA) and castor (RcLCAT-PLA) could cleave acyl chains at both the sn-1 and sn-2 positions of PC, and displayed substrate selectivity towards sn-2-ricinoleoyl-PC over sn-2-oleoyl-PC. Furthermore, co-expression of RcFAH12 with PfLCAT-PLA or RcLCAT-PLA, but not Arabidopsis AtLCAT-PLA, resulted in increased occupation of HFA at the sn-1/3 positions of TAG as well as small but insignificant increases in HFA levels in Arabidopsis seeds compared with RcFAH12 expression alone. Therefore, PfLCAT-PLA and RcLCAT-PLA may contribute to HFA turnover on PC, and represent potential candidates for engineering the production of unusual fatty acids in crops.

Cholesterol depletion sensitizes gallbladder cancer to cisplatin by impairing DNA damage response.

Gallbladder cancer (GBC) is the common malignancy of the bile tract system with extremely poor clinical outcomes, owing to its metastatic property and intrinsic resistance to the first-line drugs. Although it is well-established that cholesterol abnormity contributes to gallstone formation, a leading risk factor for GBC, the link of cholesterol homeostasis with GBC has not been investigated. The present study systematically examined the genes implicated in cholesterol homeostasis, and revealed altered gene expressions of de novo cholesterol biosynthesis and sterol sulfonation (SULT2B1), reduced bile acid synthesis (CYP7B1 and CYP39A1) and impaired sterol efflux (ABCA1, ABCG5, LCAT, and CETP) in GBC tissues. Suppression of cholesterol biosynthesis by lovastatin inhibited GBC cell proliferation possibly through attenuating the DNA repair process. Further investigation revealed lovastatin sensitized GBC cells to cisplatin-induced apoptosis and suppressed the activation of CHK1, CHK2, and H2AX during DNA damage response. By using chemically distinct statins, HMGCR depletion or supplementing mevalonate, the product of HMGCR, we showed the inhibitory effects on DNA repair process of lovastatin were due to the blockage of the mevalonate pathway. Subcutaneous xenograft mice model suggested lovastatin promoted the therapeutic efficacy of cisplatin, and significantly prolonged the survival times of tumor-bearing mice. Moreover, HMGCR ablation repressed tumor growth in vivo, which can be rescued partially by restored expression of HMGCR, suggesting the on-target effects of lovastatin. Therefore, our study provides the clinical relevance of cholesterol homeostasis with GBC progression, and highlights a novel intervention of combined use of lovastatin and cisplatin for GBC.

Arginine 123 of apolipoprotein A-I is essential for lecithin:cholesterol acyltransferase activity.

ApoA-I activates LCAT that converts lipoprotein cholesterol to cholesteryl ester (CE). Molecular dynamic simulations suggested earlier that helices 5 of two antiparallel apoA-I molecules on discoidal HDL form an amphipathic tunnel for migration of acyl chains and unesterified cholesterol to the active sites of LCAT. Our recent crystal structure of Δ(185-243)apoA-I showed the tunnel formed by helices 5/5, with two positively charged residues arginine 123 positioned at the edge of the hydrophobic tunnel. We hypothesized that these uniquely positioned residues Arg123 are poised for interaction with fatty acids produced by LCAT hydrolysis of the sn-2 chains of phosphatidylcholine, thus positioning the fatty acids for esterification to cholesterol. To test the importance of Arg123 for LCAT phospholipid hydrolysis and CE formation, we generated apoA-I[R123A] and apoA-I[R123E] mutants and made discoidal HDL with the mutants and WT apoA-I. Neither mutation of Arg123 changed the particle composition or size, or the protein conformation or stability. However, both mutations of Arg123 significantly reduced LCAT catalytic efficiency and the apparent Vmax for CE formation without affecting LCAT phospholipid hydrolysis. A control mutation, apoA-I[R131A], did not affect LCAT phospholipid hydrolysis or CE formation. These data suggest that Arg123 of apoA-I on discoidal HDL participates in LCAT-mediated cholesterol esterification.

Cellular cholesterol accumulation modulates high fat high sucrose (HFHS) diet-induced ER stress and hepatic inflammasome activation in the development of non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis (NASH), is the form of non-alcoholic fatty liver disease posing risk to progress into serious long term complications. Human and pre-clinical models implicate cellular cholesterol dysregulation playing important role in its development. Mouse model studies suggest synergism between dietary cholesterol and fat in contributing to NASH but the mechanisms remain poorly understood. Our laboratory previously reported the primary importance of hepatic endoplasmic reticulum cholesterol (ER-Chol) in regulating hepatic ER stress by comparing the responses of wild type, Ldlr-/-xLcat+/+ and Ldlr-/-xLcat-/- mice, to a 2% high cholesterol diet (HCD). Here we further investigated the roles of ER-Chol and ER stress in HFHS diet-induced NASH using the same strains. With HFHS diet feeding, both WT and Ldlr-/-xLcat+/+ accumulate ER-Chol in association with ER stress and inflammasome activation but the Ldlr-/-xLcat-/- mice are protected. By contrast, all three strains accumulate cholesterol crystal, in correlation with ER-Chol, albeit less so in Ldlr-/-xLcat-/- mice. By comparison, HCD feeding per se (i) is sufficient to promote steatosis and activate inflammasomes, and (ii) results in dramatic accumulation of cholesterol crystal which is linked to inflammasome activation in Ldlr-/-xLcat-/- mice, independent of ER-Chol. Our data suggest that both dietary fat and cholesterol each independently promote steatosis, cholesterol crystal accumulation and inflammasome activation through distinct but complementary pathways. In vitro studies using palmitate-induced hepatic steatosis in HepG2 cells confirm the key roles by cellular cholesterol in the induction of steatosis and inflammasome activations. These novel findings provide opportunities for exploring a cellular cholesterol-focused strategy for treatment of NASH.

Effect of Centella asiatica on oxidative stress and lipid metabolism in hyperlipidemic animal models.

Hyperlipidemia and many other metabolic diseases are related to oxidative stress. Centella asiatica is a traditional Chinese medicine whose antioxidant effect in vitro has been reported. We are interested in whether it possesses this effect in vivo and hence modulates lipid metabolism. Therefore, experiments were carried out on mice and golden hamsters regarding its antioxidant and hypolipidemic effect. We observed that a fraction (CAF3) of the ethanol extract (CAE) of Centella asiatica had a cholesterol decrease of 79% and a triglyceride decrease of 95% in acute mice model, so CAF3 was further investigated in high-fat-fed hamster model. It was shown that CAF3 increased SOD and GSH-Px activities and decreased MDA level, and it also improved TC, TG, LDL-C, HDL-C, AST, and ALT levels. L-CAT and SR-BI gene expression in hamsters were increased. Taken together, our data suggest that the CAF3 fraction of Centella asiatica has antioxidant and hypolipidemic properties.

Scavenger receptor BI and high-density lipoprotein regulate thymocyte apoptosis in sepsis.

OBJECTIVE: Thymocyte apoptosis is a major event in sepsis; however, how this process is regulated remains poorly understood. APPROACH AND RESULTS: Septic stress induces glucocorticoids production which triggers thymocyte apoptosis. Here, we used scavenger receptor BI (SR-BI)-null mice, which are completely deficient in inducible glucocorticoids in sepsis, to investigate the regulation of thymocyte apoptosis in sepsis. Cecal ligation and puncture induced profound thymocyte apoptosis in SR-BI(+/+) mice, but no thymocyte apoptosis in SR-BI(-/-) mice because of lack of inducible glucocorticoids. Unexpectedly, supplementation of glucocorticoids only partly restored thymocyte apoptosis in SR-BI(-/-) mice. We demonstrated that high-density lipoprotein (HDL) is a critical modulator for thymocyte apoptosis. SR-BI(+/+) HDL significantly enhanced glucocorticoid-induced thymocyte apoptosis, but SR-BI(-/-) HDL had no such activity. Further study revealed that SR-BI(+/+) HDL modulates glucocorticoid-induced thymocyte apoptosis via promoting glucocorticoid receptor translocation, but SR-BI(-/-) HDL loses such regulatory activity. To understand why SR-BI(-/-) HDL loses its regulatory activity, we analyzed HDL cholesterol contents. There was 3-fold enrichment of unesterified cholesterol in SR-BI(-/-) HDL compared with SR-BI(+/+) HDL. Normalization of unesterified cholesterol in SR-BI(-/-) HDL by probucol administration or lecithin cholesteryl acyltransferase expression restored glucocorticoid-induced thymocyte apoptosis, and incorporating unesterified cholesterol into SR-BI(+/+) HDL rendered SR-BI(+/+) HDL dysfunctional. Using lckCre-GR(fl/fl) mice in which thymocytes lack cecal ligation and puncture-induced thymocyte apoptosis, we showed that lckCre-GR(fl/fl) mice were significantly more susceptible to cecal ligation and puncture-induced septic death than GR(fl/fl) control mice, suggesting that glucocorticoid-induced thymocyte apoptosis is required for protection against sepsis. CONCLUSIONS: The findings in this study reveal a novel regulatory mechanism of thymocyte apoptosis in sepsis by SR-BI and HDL.

Lecithin/cholesterol acyltransferase modulates diet-induced hepatic deposition of triglycerides in mice.

Lecithin/cholesterol acyltransferase (LCAT) is responsible for the esterification of the free cholesterol of plasma lipoproteins. Here, we investigated the involvement of LCAT in mechanisms associated with diet-induced hepatic triglyceride accumulation in mice. LCAT-deficient (LCAT(-/-)) and control C57BL/6 mice were placed on a Western-type diet (17.3% protein, 48.5% carbohydrate, 21.2% fat, 0.2% cholesterol, 4.5kcal/g) for 24weeks, then histopathological and biochemical analyses were performed. We report that, in our experimental setup, male LCAT(-/-) mice are characterized by increased diet-induced hepatic triglyceride deposition and impaired hepatic histology and architecture. Mechanistic analyses indicated that LCAT deficiency was associated with enhanced intestinal absorption of dietary triglycerides (3.6±0.5mg/dl per minute for LCAT(-/-) vs. 2.0±0.7mg/dl per minute for C57BL/6 mice; P<.05), accelerated clearance of postprandial triglycerides and a reduced rate of hepatic very low density lipoprotein triglyceride secretion (9.8±1.1mg/dl per minute for LCAT(-/-) vs. 12.5±1.3mg/dl per minute for C57BL/6 mice, P<.05). No statistical difference in the average daily food consumption between mouse strains was observed. Adenovirus-mediated gene transfer of LCAT in LCAT(-/-) mice that were fed a Western-type diet for 12weeks resulted in a significant reduction in hepatic triglyceride content (121.2±5.9mg/g for control infected mice vs. 95.1±5.8mg/g for mice infected with Ad-LCAT, P<.05) and a great improvement of hepatic histology and architecture. Our data extend the current knowledge on the functions of LCAT, indicating that LCAT activity is an important modulator of processes associated with diet-induced hepatic lipid deposition.

High hydrostatic pressure extract of garlic increases the HDL cholesterol level via up-regulation of apolipoprotein A-I gene expression in rats fed a high-fat diet.

BACKGROUND: Cardiovascular disease (CVD) is the number one cause of mortality worldwide and a low high-density lipoprotein cholesterol (HDL-C) level is an important marker of CVD risk. Garlic (Allium sativum) has been widely used in the clinic for treatment of CVD and regulation of lipid metabolism. This study investigated the effects of a high hydrostatic pressure extract of garlic (HEG) on HDL-C level and regulation of hepatic apolipoprotein A-I (apoA-I) gene expression. METHODS: Male Sprague-Dawley rats were divided into two groups and maintained on a high-fat control diet (CON) or high-fat control diet supplemented with high hydrostatic pressure extract of garlic (HEG) for 5 weeks. Changes in the expression of genes related to HDL-C metabolism were analyzed in liver, together with biometric and blood parameters. RESULTS: In the HEG group, the plasma triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) levels were significantly decreased in comparison with the CON group (P < 0.05). Dietary HEG also lowered the hepatic TG and total cholesterol (TC) levels compared to the CON group. While the plasma HDL-C level and mRNA level of hepatic apoA-I, which is one of primarily proteins of HDL-C particle, were significantly increased in the HEG group compared to the CON group (P < 0.05). The gene expression of ATP-binding cassette transporter A1 (ABCA1) and lecithin:cholesterol acyltransferase (LCAT), importantly involved in the biogenesis in HDL, were also up-regulated by dietary HEG. CONCLUSIONS: These results suggest that HEG ameliorates plasma lipid profiles and attenuates hepatic lipid accumulation in the high-fat fed rats. Our findings provides that the effects of HEG on the increase of the plasma HDL-C level was at least partially mediated by up-regulation of hepatic genes expression such as apoA-I, ABCA1, and LCAT in rats fed a high-fat diet.

Hypolipidemic effects of a new piperine derivative GB-N from Piper longum in high-fat diet-fed rats.

CONTEXT: Long pepper, Piper longum Linn. (Piperaceae), is widely used in traditional Mongolian medicine for treating hyperlipidemia and coronary heart disease. OBJECTIVE: To investigate the hypolipidemic effects of a new piperine derivative GB-N isolated from long pepper in high-fat diet-fed rats. METHODS: The levels of serum total cholesterol, triacylglycerols (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were determined by enzymatic colorimetric method. The levels of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA), CYP7A1, lecithin cholesterol acyltransferase (LCAT) and LDL receptor (LDLR) mRNA and protein expression were detected by real-time polymerase chain reaction and western blot analysis. RESULTS AND DISCUSSION: Compared with model rats, oral administration of GB-N at doses of 2.5-10 mg/kg to hyperlipidemic rats could significantly decrease the levels of serum TG from 1.54 mmol/L in hyperlipidemic rats to 0.94-1.02 mmol/L, with an increase in serum HDL-C levels from 0.40 mmol/L in hyperlipidemic rats to 1.21-2.26 mmol/L. Treatment with GB-N (10 mg/kg) could also significantly upregulate levels of hepatic HMG-CoA reductase, CYP7A1, LCAT and LDLR mRNA and protein expression. CONCLUSION: GB-N had hypolipidemic activity via regulating lipid metabolism pathways in liver of hyperlipidemic rats and could be explored as a potential agent for the prevention of hyperlipidemia diseases.

Regulation of the expression of key genes involved in HDL metabolism by unsaturated fatty acids.

The cardioprotective effects of HDL have been largely attributed to their role in the reverse cholesterol transport pathway, whose efficiency is affected by many proteins involved in the formation and remodelling of HDL. The aim of the present study was to determine the effects, and possible mechanisms of action, of unsaturated fatty acids on the expression of genes involved in HDL metabolism in HepG2 cells. The mRNA concentration of target genes was assessed by real-time PCR. Protein concentrations were determined by Western blot or immunoassays. PPAR and liver X receptor (LXR) activities were assessed in transfection experiments. Compared with the SFA palmitic acid (PA), the PUFA arachidonic acid (AA), EPA and DHA significantly decreased apoA-I, ATP-binding cassette A1 (ABCA1), lecithin-cholesterol acyltransferase (LCAT) and phospholipid transfer protein mRNA levels. EPA and DHA significantly lowered the protein concentration of apoA-I and LCAT in the media, as well as the cellular ABCA1 protein content. In addition, DHA repressed the apoA-I promoter activity. AA lowered only the protein concentration of LCAT in the media. The activity of PPAR was increased by DHA, while the activity of LXR was lowered by both DHA and AA, relative to PA. The regulation of these transcription factors by PUFA may explain some of the PUFA effects on gene expression. The observed n-3 PUFA-mediated changes in gene expression are predicted to reduce the rate of HDL particle formation and maturation.

Lecithin cholesterol acyltransferase: an anti- or pro-atherogenic factor?

Lecithin cholesterol acyl transferase (LCAT) is a plasma enzyme that esterifies cholesterol and raises high-density lipoprotein cholesterol, but its role in atherosclerosis is not clearly established. Studies of various animal models have yielded conflicting results, but studies done in rabbits and non-human primates, which more closely simulate human lipoprotein metabolism, indicate that LCAT is likely atheroprotective. Although suggestive, there are also no biomarker studies that mechanistically link LCAT with cardiovascular disease. Imaging studies of patients with LCAT deficiency have also not yielded a clear answer to the role of LCAT in atherosclerosis. Recombinant LCAT, however, is currently being developed as a therapeutic product for enzyme replacement therapy of patients with genetic disorders of LCAT for the prevention and/or treatment of renal disease, but it may also have value for the treatment of acute coronary syndrome.

Dietary capsanthin, the main carotenoid in paprika (Capsicum annuum), alters plasma high-density lipoprotein-cholesterol levels and hepatic gene expression in rats.

The effects of dietary capsanthin, the main carotenoid in paprika (Capsicum annuum), on lipid metabolism were examined. Young male Wistar rats were fed diets containing paprika powder, paprika organic solvent extract, residue of paprika extract, and purified capsanthin. Administration of purified capsanthin for 2 weeks resulted in a significant increase in plasma HDL-cholesterol (P < 0.05) without detectable differences in plasma total cholesterol and TAG concentrations. A statistically significant correlation (r 0.567; P < 0.001) was found between dietary capsanthin concentrations and plasma HDL-cholesterol concentrations. Animals receiving diets containing two different capsanthin concentrations exhibited dose-dependent increases in plasma HDL-cholesterol (r 0.597; P < 0.005). While capsanthin was absent in the liver of animals fed the basal diet, it increased markedly in capsanthin-fed animals (P < 0.001). Quantitative analyses of hepatic mRNA levels revealed that capsanthin administration resulted in up-regulation of mRNA for apoA5 and lecithin cholesterol acyltransferase (LCAT), without significant differences in other mRNA levels related to HDL-cholesterol metabolism. These results suggest that capsanthin had an HDL-cholesterol-raising effect on plasma, and the potential to increase cholesterol efflux to HDL particles by increasing apoA5 levels and/or enhancement of LCAT activity.

Compound heterozygosity (G71R/R140H) in the lecithin:cholesterol acyltransferase (LCAT) gene results in an intermediate phenotype between LCAT-deficiency and fish-eye disease.

The esterification of free cholesterol (FC) in plasma, catalyzed by the enzyme lecithin:cholesterol acyltransferase (LCAT; EC 2.3.1.43), is a key process in lipoprotein metabolism. The resulting cholesteryl esters (CE) represent the main core lipids of low (LDL) and high density lipoproteins (HDL). Primary (familial) LCAT-deficiency (FLD) is a rare autosomal recessive genetic disease caused by the complete or near absence of LCAT activity. In fish-eye disease (FED), residual LCAT activity is still detectable. Here, we describe a 32-year-old patient with corneal opacity, very low LCAT activity, reduced amounts of CE (low HDL-cholesterol level), and elevated triglyceride (TG) values. The lipoprotein pattern was abnormal with regard to lipoprotein composition and concentration, but distinct lipoprotein classes were still present. Despite of typical features of glomerular proteinuria, creatinine clearance was normal. DNA sequencing and restiction fragment analyses revealed two separate mutations in the patient's LCAT gene: a previously described G to A transition in exon 4 converting Arg140 to His, inherited from his mother, and a novel G to C transversion in exon 2 converting Gly71 to Arg, inherited from his father, indicating that M.P. was a compound heterozygote. Determination of enzyme activities of recombinant LCAT proteins obtained upon transfection of COS-7 cells with plasmids containing G71R-LCAT or wild-type LCAT cDNA revealed very low alpha- and absence of beta-LCAT activity for the G71R mutant. The identification of the novel G71R LCAT mutation supports the proposed molecular model for the enzyme implying that the "lid" domain at residues 50-74 is involved in enzyme:substrate interaction. Our data are in line with the hypothesis that a key event in the etiology of FLD is the loss of distinct lipoprotein fractions.

Dietary fats differentially modulate the expression of lecithin:cholesterol acyltransferase, apoprotein-A1 and scavenger receptor b1 in rats.

In the present study the effects of dietary fat with defined fatty acids on lecithin:cholesterol acyltransferase (LCAT) and apoA-1, the two components of HDL that play a major role in reverse cholesterol transport (RCT), were examined. In addition, the expression of scavenger receptor B1 (SR-B1), the receptor involved in the uptake of HDL core lipids, was also determined under the same conditions in rats fed semisynthetic diets supplemented with triolein (TO), tripalmitin (TP) or menhaden oil (MO). Serum LCAT activity [ micro mol CE/(L.h)] was significantly (P < 0.05) higher in rats fed TO (33 +/- 4) compared with those fed TP (23 +/- 3) or MO (21 +/- 1). The levels of hepatic LCAT mRNA and hepatic SR-B1 receptor protein did not differ between rats fed TP and MO. The triolein diet, on the other hand, increased the induction of hepatic LCAT mRNA and hepatic SR-B1 receptor protein 1.5- to 2-fold. Serum HDL cholesterol concentrations differed among all groups and were 1.30 +/- 0.08, 1.17 +/- 0.10 and 0.91 +/- 0.06 mmol/L for TO-, TP- and MO-fed rats, respectively. Serum apoA-1 levels were significantly higher in TO-fed rats than in the other two groups. The data indicate that TO increases the secretion of HDL and its components (apoA-1 and LCAT), and stimulates the production of hepatic SR-B1 receptor protein. Overall, these results suggest that triolein may promote RCT and thus retard the development of atherosclerosis.

Analysis of gene expression following spinal cord injury in rat using complementary DNA microarray.

To identify genes that were altered by spinal cord injury (SCI), we used complementary DNA microarray consisting 1176 rat genes. Rats were subjected to contusive injury of the thoracic spinal cord. Sham animals received only a laminectomy. Twenty-four hours later, spinal cord was dissected out, a 32P labeled probe was prepared and hybridized to the microarray. We identified three genes that showed a greater than 2-fold increase in SCI tissue, heat shock 27-kDa protein, tissue inhibitor of metalloproteinase-1 and epidermal fatty acid-binding protein. Seven genes, lecithin:cholesterol acyltransferase, dipeptidyl aminopeptidase related protein, phospholipase C delta 4, plasma membrane Ca2+-ATPase isoform 2, G-protein GO alpha subunit, GABA transporter 3, and neuroendrocrine protein 7B2 were down-regulated greater than 50% in SCI tissue. Changes in expression of these genes were confirmed by reverse transcription-polymerase chain reaction. These genes may play a role in the response to tissue damage or repair following SCI.

[Analysis of lecithin-cholesterol acyltransferase cDNA and protein sequence from tree shrew].

OBJECTIVE: To acquire cDNA sequence of lecithin-cholesterol acyltransferase (LCAT) from tree shrew and analyze the sequence structure. METHODS: The first strand cDNA was acquired by reverse transcription using mRNA from tree shrew liver as template. By the method of SMART RACE PCR, tree shrew LCAT cDNA was acquired and deduced its amino acids sequence. The sequence and structure of tree shrew LCAT cDNA and amino acid were analyzed and predicted by the molecular software. RESULTS: Tree shrew LCAT cDNA is composed of 1,340 bp, including 2 bp 5' untranslated region (5' UTR), 1,320 bp open reading frame (ORF) which encodes protein precursor of 440 amino acids (24 amino acids signal peptide and 416 amino acids mature peptide), and 18 bp 3' untranslated region (3'UTR). The stop codon is TAA and there is a poly (A) signal sequence AATAAA and a 25 bp poly (A) tail. Tree shrew LCAT cDNA sequence has been accepted by GenBank as a new gene, accession number AF272861 and its homology with human and baboon was 90% and 89%, respectively. CONCLUSION: The sequence of LCAT cDNA in tree shrew has high identity with that of human and other experimental animal species.

Evolutionary history of the most speciose mammals: molecular phylogeny of muroid rodents.

Phylogenetic relationships between 32 species of rodents representing 14 subfamilies of Muridae and four subfamilies of Dipodidae were studied using sequences of the nuclear protein-coding genes Lecithin Cholesterol Acyl Transferase (LCAT) and von Willebrand Factor (vWF). An examination of some evolutionary properties of each data matrix indicates that the two genes are rather complementary, with lower rates of nonsynonymous substitutions for LCAT. Both markers exhibit a wide range of GC3 percentages (55%-89%), with several taxa above 70% GC3 for vWF, which indicates that those exonic regions might belong to the richest class of isochores. The primary sequence data apparently harbor few saturations, except for transitions on third codon positions for vWF, as indicated by comparisons of observed and expected pairwise values of substitutions. Phylogenetic trees based on 1,962 nucleotidic sites from the two genes indicate that the 14 Muridae subfamilies are organized into five major lineages. An early isolation leads to the clade uniting the fossorial Spalacinae and semifossorial Rhizomyinae with a strong robustness. The second lineage includes a series of African taxa representing nesomyines, dendromurines, cricetomyines, and the sole living member of mystromyines. The third one comprises only the mouselike hamster CALOMYSCUS: The fourth clade represents the cricetines, myospalacines, sigmodontines, and arvicolines, whereas the fifth one comprises four "traditional" subfamilies (Gerbillinae, Murinae, Otomyinae, and Acomyinae). Within these groups, we confirm the monophyly of almost all studied subfamilies, namely, Spalacinae, Rhizomyinae, Nesomyinae, Cricetomyinae, Arvicolinae, Sigmodontinae, Cricetinae, Gerbillinae, Acomyinae, and Murinae. Finally, we present evidence that the sister group of Acomyinae is Gerbillinae, and we confirm a nested position of Myospalacinae within Cricetinae and Otomyinae within Murinae. From a biogeographical point of view, the five main lineages spread and radiated from Asia with different degrees of success: the first three groups are now represented by a limited number of species and genera localized in some regions, whereas the last two groups radiated in a large variety of species and genera dispersed all over the world.

Linkage of a candidate gene locus to familial combined hyperlipidemia: lecithin:cholesterol acyltransferase on 16q.

Familial combined hyperlipidemia (FCHL) is a common lipid disorder characterized by elevated levels of plasma cholesterol and triglycerides that is present in 10% to 20% of patients with premature coronary artery disease. To study the pathophysiological basis and genetics of FCHL, we previously reported recruitment of 18 large families. We now report linkage studies of 14 candidate genes selected for their potential involvement in the aspects of lipid and lipoprotein metabolism that are altered in FCHL. We used highly polymorphic markers linked to the candidate genes, and these markers were analyzed using several complementary, nonparametric statistical allele-sharing linkage methodologies. This current sample has been extended over the one in which we identified an association with the apolipoprotein (apo) AI-CIII-AIV gene cluster. We observed evidence for linkage of this region and FCHL (P<0.001), providing additional support for its involvement in FCHL. We also identified a new locus showing significant evidence of linkage to the disorder: the lecithin:cholesterol acyltransferase (LCAT) locus (P<0.0006) on chromosome 16. In addition, analysis of the manganese superoxide dismutase locus on chromosome 6 revealed a suggestive linkage result in this sample (P<0.006). Quantitative traits related to FCHL also provided some evidence of linkage to these regions. No evidence of linkage to the lipoprotein lipase gene, the microsomal triglyceride transfer protein gene, or several other genes involved in lipid metabolism was observed. The data suggest that the lecithin:cholesterol acyltransferase and apolipoprotein AI-CIII-AIV loci may act as modifying genes contributing to the expression of FCHL.