A Vaccine Targeted at CETP Alleviates High Fat and High Cholesterol Diet-Induced Atherosclerosis and Non-Alcoholic Steatohepatitis in Rabbit

Low HDL-C levels are associated with atherosclerosis and non-alcoholic steatohepatitis, and increased levels may reduce the risk of these diseases. Inhibition of cholesteryl ester transfer protein (CETP) activity is considered a promising strategy for increasing HDL-C levels. Since CETP is a self-antigen with low immunogenicity, we developed a novel CETP vaccine (Fc-CETP₆) to overcome the low immunogenicity of CETP and for long-term inhibition of CETP activity. The vaccine consists of a rabbit IgG Fc domain for antigen delivery to antigen-presenting cells fused to a linear array of 6 repeats of a CETP epitope to efficiently activate B cells. Rabbits were fed a high fat/cholesterol (HFC) diet to induce atherosclerosis and NASH, and immunized with Fc-CETP₆ vaccine. The Fc-CETP₆ vaccine successfully elicited anti-CETP antibodies and lowered plasma CETP activity. The levels of plasma HDL-C and ApoA-I were higher, and plasma ox-LDL lower, in the Fc-CETP₆-immunized rabbits as compared to the unimmunized HFC diet-fed rabbits. Pathological analyses revealed less lipid accumulation and inflammation in the aorta and liver of the Fc-CETP₆-immunized rabbits. These results show that the Fc-CETP₆ vaccine efficiently elicited antibodies against CETP and reduced susceptibility to both atherosclerosis and steatohepatitis induced by the HFC diet. Our findings suggest that the Fc-CETP₆ vaccine may improve atherosclerosis and NASH and has high potential for clinical use.     

Effect of apoA-I Mutations in the Capacity of Reconstituted HDL to Promote ABCG1-Mediated Cholesterol Efflux

ATP binding cassette transporter G1 (ABCG1) mediates the cholesterol transport from cells to high-density lipoprotein (HDL), but the role of apolipoprotein A-I (apoA-I), the main protein constituent of HDL, in this process is not clear. To address this, we measured cholesterol efflux from HEK293 cells or J774 mouse macrophages overexpressing ABCG1 using as acceptors reconstituted HDL (rHDL) containing wild-type or various mutant apoA-I forms. It was found that ABCG1-mediated cholesterol efflux was severely reduced (by 89%) when using rHDL containing the carboxyl-terminal deletion mutant apoA-I[Δ(185–243)]. ABCG1-mediated cholesterol efflux was not affected or moderately decreased by rHDL containing amino-terminal deletion mutants and several mid-region deletion or point apoA-I mutants, and was restored to 69–99% of control by double deletion mutants apoA-I[Δ(1–41)Δ(185–243)] and apoA-I[Δ(1–59)Δ(185–243)]. These findings suggest that the central helices alone of apoA-I associated to rHDL can promote ABCG1-mediated cholesterol efflux. Further analysis showed that rHDL containing the carboxyl-terminal deletion mutant apoA-I[Δ(185–243)] only slightly reduced (by 22%) the ABCG1-mediated efflux of 7-ketocholesterol, indicating that depending on the sterol type, structural changes in rHDL-associated apoA-I affect differently the ABCG1-mediated efflux of cholesterol and 7-ketocholesterol. Overall, our findings demonstrate that rHDL-associated apoA-I structural changes affect the capacity of rHDL to accept cellular cholesterol by an ABCG1-mediated process. The structure-function relationship seen here between rHDL-associated apoA-I mutants and ABCG1-mediated cholesterol efflux closely resembles that seen before in lipid-free apoA-I mutants and ABCA1-dependent cholesterol efflux, suggesting that both processes depend on the same structural determinants of apoA-I.     

Evidence for a role of CETP in HDL remodeling and cholesterol efflux: Role of cysteine 13 of CETP

Cholesteryl ester transfer protein (CETP), a key regulator of high-density lipoprotein (HDL) metabolism, induces HDL remodeling by transferring lipids between apolipoprotein B-containing lipoproteins and HDL, and/or by promoting lipid transfer between HDL subparticles. In this study, we investigated the mechanism as to how CETP induces the generation of lipid-poor particles (pre-β-HDL) from HDL, which increases ATP-binding cassette transporter 1-mediated cholesterol efflux. This CETP-dependent HDL remodeling is enhanced by the CETP modulator dalcetrapib both in plasma and isolated HDL. The interaction of dalcetrapib with cysteine 13 of CETP is required, since this effect was abolished when using mutant CETP in which cysteine 13 was substituted for a serine residue. Other thiol-containing compounds were identified as CETP modulators interacting with cysteine 13 of CETP. In order to mimic dalcetrapib-bound CETP, mutant CETP proteins were prepared by replacing cysteine 13 with the bulky amino acid tyrosine or tryptophan. The resultant mutants showed virtually no CETP-dependent lipid transfer activity but demonstrated preserved CETP-dependent pre-β-HDL generation. Overall, these data demonstrate that the two functions of CETP i.e., cholesteryl ester transfer and HDL remodeling can be uncoupled by interaction of thiol-containing compounds with cysteine 13 of CETP or by introducing large amino acid residues in place of cysteine 13.     

Remodeling of HDL by CETP in vivo and by CETP and hepatic lipase in vitro results in enhanced uptake of HDL CE by cells expressing scavenger receptor B-I.

The transport of HDL cholesteryl esters (CE) from plasma to the liver involves a direct uptake pathway, mediated by hepatic scavenger receptor B-I (SR-BI), and an indirect pathway, involving the exchange of HDL CE for triglycerides (TG) of TG-rich lipoproteins by cholesteryl ester transfer protein (CETP). We carried out HDL CE turnover studies in mice expressing human CETP and/or human lecithin:cholesterol acyltransferase (LCAT) transgenes on a background of human apoA-I expression. The fractional clearance of HDL CE by the liver was delayed by LCAT transgene, while the CETP transgene increased it. However, there was no incremental transfer of HDL CE radioactivity to the TG-rich lipoprotein fraction in mice expressing CETP, suggesting increased direct removal of HDL CE in the liver. To evaluate the possibility that this might be mediated by SR-BI, HDL isolated from plasma of the different groups of transgenic mice was incubated with SR-BI transfected or control CHO cells. HDL isolated from mice expressing CETP showed a 2- to 4-fold increase in SR-BI-mediated HDL CE uptake, compared to HDL from mice lacking CETP. The addition of pure CETP to HDL in cell culture did not lead to increased selective uptake of HDL CE by cells. However, when human HDL was enriched with TG by incubation with TG-rich lipoproteins in the presence of CETP, then treated with hepatic lipase, there was a significant enhancement of HDL CE uptake. Thus, the remodeling of human HDL by CETP, involving CE;-TG interchange, followed by the action of hepatic lipase (HL), leads to the enhanced uptake of HDL CE by cellular SR-BI.These observations suggest that in animals such as humans in which both the selective uptake and CETP pathways are active, the two pathways could operate in a synergistic fashion to enhance reverse cholesterol transport.     

Intact Cohesion,Anaphase, and Chromosome Segregation in Human Cells Harboring Tumor-Derived Mutations in STAG2

Somatic mutations of the cohesin complex subunit STAG2 are present in diverse tumor types. We and others have shown that STAG2 inactivation can lead to loss of sister chromatid cohesion and alterations in chromosome copy number in experimental systems. However, studies of naturally occurring human tumors have demonstrated little, if any, correlation between STAG2 mutational status and aneuploidy, and have further shown that STAG2-deficient tumors are often euploid. In an effort to provide insight into these discrepancies, here we analyze the effect of tumor-derived STAG2 mutations on the protein composition of cohesin and the expected mitotic phenotypes of STAG2 mutation. We find that many mutant STAG2 proteins retain their ability to interact with cohesin; however, the presence of mutant STAG2 resulted in a reduction in the ability of regulatory subunits WAPL, PDS5A, and PDS5B to interact with the core cohesin ring. Using AAV-mediated gene targeting, we then introduced nine tumor-derived mutations into the endogenous allele of STAG2 in cultured human cells. While all nonsense mutations led to defects in sister chromatid cohesion and a subset induced anaphase defects, missense mutations behaved like wild-type in these assays. Furthermore, only one of nine tumor-derived mutations tested induced overt alterations in chromosome counts. These data indicate that not all tumor-derived STAG2 mutations confer defects in cohesion, chromosome segregation, and ploidy, suggesting that there are likely to be other functional effects of STAG2 inactivation in human cancer cells that are relevant to cancer pathogenesis.     

Novel ATPase Cu2+ Transporting Beta Polypeptide Mutations in Chinese Families with Wilson's Disease

Wilson''s disease (WD) is an autosomal recessive inherited disorder caused by mutations in the ATPase Cu²⁺ transporting beta polypeptide gene (ATP7B). The detailed metabolism of copper-induced pathology in WD is still unknown. Gene mutations as well as the possible pathways involved in the ATP7B deficiency were documented. The ATP7B gene was analyzed for mutations in 18 Chinese Han families with WD by direct sequencing. Cell viability and apoptosis analysis of ATP7B small interfering RNA (siRNA)-treated human liver carcinoma (HepG2) cells were measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and Hoechst 33342 staining. Finally, the expression of B-cell CLL/lymphoma 2 (BCL2), BCL2-associated X protein (BAX), sterol regulatory element binding protein 1 (SREBP1), and minichromosome maintenance protein 7 (MCM7) of ATP7B siRNA-treated cells were tested by real-time polymerase chain reaction (real-time PCR) and Western blot analysis. Twenty different mutations including four novel mutations (p.Val145Phe, p.Glu388X, p.Thr498Ser and p.Gly837X) in the ATP7B gene were identified in our families. Haplotype analysis revealed that founder effects for four mutations (p.Arg778Leu, p.Pro992Leu, p.Ile1148Thr and p.Ala1295Val) existed in these families. Transfection of HepG2 cells with ATP7B siRNA resulted in decreased mRNA expression by 86.3%, 93.1% and 90.8%, and decreased protein levels by 58.5%, 85.5% and 82.1% at 24, 48 and 72 hours, respectively (All P<0.01). In vitro study revealed that the apoptotic, cell cycle and lipid metabolism pathway may be involved in the mechanism of WD. Our results revealed that the genetic cause of 18 Chinese families with WD and ATP7B deficiency-induce apoptosis may result from imbalance in cell cycle and lipid metabolism pathway.     

Association Between Lipids, Lipoproteins Composition of HDL Particles and Triglyceride-Rich Lipoproteins, and LCAT and CETP Activity in Post-renal Transplant Patients

High-density lipoprotein (HDL) remodeling within the plasma compartment and the association between lecithin-cholesterol acyltransferase (LCAT) and cholesterol ester transfer protein (CETP) activity, and lipid, lipoprotein concentrations and composition were investigated. The aim was to examine the high sensitivity of C-reactive protein (hsCRP), lipid, apolipoprotein B (apoB), apoAI, total apoAII, apoAIInonB, apoB-containing apoAII (apoB:AII), total apoCIII, apoCIIInonB, apoB-containing apoCIII (apoB:CIII) concentration and LCAT and CETP activity to gain an insight into the association between them and LCAT and CETP, 57 post-renal transplant (Tx) patients with and without statin therapy and in 15 healthy subjects. Tx patients had moderate hypertriglyceridemia, hypercholesterolemia, and dyslipoproteinemia, disturbed triglyceride-rich lipoproteins (TRLs) and HDL composition, decreased LCAT, and slightly increased hsCRP but no CETP activity. Spearman’s correlation test showed the association between lipids and lipoproteins and LCAT or CETP, and multiple ridge stepwise forward regression showed that immunosuppressive therapy in Tx patients can disturb HDL and TRLs composition. The results suggest that inhibition or activation of LCAT is due, in part, to HDL-associated lipoprotein. Lipoprotein composition of apoAI, apoAIInonB, and apoCIIInonB in HDL particle and apoB:AII TRLs can contribute to decrease LCAT mass in Tx patients. Tx patients without statin and with lower triglycerides but higher HDL cholesterol concentration and disturbed lipoprotein composition of ApoAI and apoAII in HDL particle can decrease LCAT, increase LDL cholesterol, aggravate renal graft, and accelerate atherosclerosis and chronic heart diseases.     

Segregation of a Latent High Adiposity Phenotype in Families with a History of Type 2 Diabetes Mellitus Implicates Rare Obesity-Susceptibility Genetic Variants with Large Effects in Diabetes-Related Obesity

Background

We recently reported significantly greater weight gain in non-diabetic healthy subjects with a 1^st degree family history (FH+) of type 2 diabetes mellitus (T2DM) than in a matched control group without such history (FH−) during voluntary overfeeding, implying co-inheritance of susceptibilities to T2DM and obesity. We have estimated the extent and mode of inheritance of susceptibility to increased adiposity in FH+.

Methods

Normoglycaemic participants were categorised either FH+ (≥1 1^st degree relative with T2DM, 50F/30M, age 45±14 (SD) yr) or FH− (71F/51M, age 43±14 yr). Log-transformed anthropometric measurements (height, hip and waist circumferences) and lean, bone and fat mass (Dual Energy X-ray Absorptiometry) data were analysed by rotated Factor Analysis. The age- and gender-adjusted distributions of indices of adiposity in FH+ were assessed by fits to a bimodal model and by relative risk ratios (RR, FH+/FH−) and interpreted in a purely genetic model of FH effects.

Results

The two orthogonal factors extracted, interpretable as Frame and Adiposity accounted for 80% of the variance in the input data. FH+ was associated with significantly higher Adiposity scores (p<0.01) without affecting Frame scores. Adiposity scores in FH+ conformed to a bimodal normal distribution, consistent with dominant expression of major susceptibility genes with 59% (95% CI 40%, 74%) of individuals under the higher mode. Calculated risk allele frequencies were 0.09 (0.02, 0.23) in FH−, 0.36 (0.22, 0.48) in FH+ and 0.62 (0.36, 0.88) in unobserved T2DM-affected family members.

Conclusions

The segregation of Adiposity in T2DM-affected families is consistent with dominant expression of rare risk variants with major effects, which are expressed in over half of FH+ and which can account for most T2DM-associated obesity in our population. The calculated risk allele frequency in FH− suggests that rare genetic variants could also account for a substantial fraction of the prevalent obesity in this society.     

Telomere Length and the Risk of Cutaneous Malignant Melanoma in Melanoma-Prone Families with and without CDKN2A Mutations

Introduction

Recent evidence suggests a link between constitutional telomere length (TL) and cancer risk. Previous studies have suggested that longer telomeres were associated with an increased risk of melanoma and larger size and number of nevi. The goal of this study was to examine whether TL modified the risk of melanoma in melanoma-prone families with and without CDKN2A germline mutations.

Materials and Methods

We measured TL in blood DNA in 119 cutaneous malignant melanoma (CMM) cases and 208 unaffected individuals. We also genotyped 13 tagging SNPs in TERT.

Results

We found that longer telomeres were associated with an increased risk of CMM (adjusted OR = 2.81, 95% CI = 1.02–7.72, P = 0.04). The association of longer TL with CMM risk was seen in CDKN2A- cases but not in CDKN2A+ cases. Among CMM cases, the presence of solar injury was associated with shorter telomeres (P = 0.002). One SNP in TERT, rs2735940, was significantly associated with TL (P = 0.002) after Bonferroni correction.

Discussion

Our findings suggest that TL regulation could be variable by CDKN2A mutation status, sun exposure, and pigmentation phenotype. Therefore, TL measurement alone may not be a good marker for predicting CMM risk.     

Novel ENAM and LAMB3 Mutations in Chinese Families with Hypoplastic Amelogenesis Imperfecta

Amelogenesis imperfecta is a group of inherited diseases affecting the quality and quantity of dental enamel. To date, mutations in more than ten genes have been associated with non-syndromic amelogenesis imperfecta (AI). Among these, ENAM and LAMB3 mutations are known to be parts of the etiology of hypoplastic AI in human cases. When both alleles of LAMB3 are defective, it could cause junctional epidermolysis bullosa (JEB), while with only one mutant allele in the C-terminus of LAMB3, it could result in severe hypoplastic AI without skin fragility. We enrolled three Chinese families with hypoplastic autosomal-dominant AI. Despite the diagnosis falling into the same type, the characteristics of their enamel hypoplasia were different. Screening of ENAM and LAMB3 genes was performed by direct sequencing of genomic DNA from blood samples. Disease-causing mutations were identified and perfectly segregated with the enamel defects in three families: a 19-bp insertion mutation in the exon 7 of ENAM (c.406_407insTCAAAAAAGCCGACCACAA, p.K136Ifs*16) in Family 1, a single-base deletion mutation in the exon 5 of ENAM (c. 139delA, p. M47Cfs*11) in Family 2, and a LAMB3 nonsense mutation in the last exon (c.3466C>T, p.Q1156X) in Family 3. Our results suggest that heterozygous mutations in ENAM and LAMB3 genes can cause hypoplastic AI with markedly different phenotypes in Chinese patients. And these findings extend the mutation spectrum of both genes and can be used for mutation screening of AI in the Chinese population.     

Germline Mutations of Tetranucleotide DNA Repeats in Families with Normal Children and Reproductive Pathology

We have previously reported a high rate of tetranucleotide DNA repeat mutations, including mutations of both germline and somatic origin, in spontaneous human abortions. To analyze in more detail mutational microsatellite (MS) variability in meiosis and its possible association with disturbed embryonic development, we have conducted a comparative study of mutation rates of a panel of 15 autosomal tetranucleotide MSs in 55 families with healthy children and in 103 families that have had spontaneous abortions with normal karyotypes. In the families with miscarriage, the gametic MS mutation rate was higher than in the families with normal reproductive function (4.36 × 10⁻³ versus 2.32 × 10⁻³ per locus per gamete per generation), but this difference was statistically nonsignificant (P = 0.25). No association of MS mutations with familiar miscarriage was found. Mutations at the MS loci studied were recorded almost 3 times as often in spermatogenesis as in oogenesis, which is likely to result from a greater number of DNA replication cycles in male germline cell precursors than in female ones. Mutations increasing and reducing the MS sequence length appeared at virtually the same rate. Changes in MS DNA sequence length per one repeated element, i.e., single-step mutations (93% of cases) exceeded all other events of allele length change. The highest number of mutations (81.2%) was found in longer alleles. This distribution of mutations by size, direction, and parental origin corresponds to the multistep mutation model of their emergence via mechanism of DNA strand slippage during replication.__________Translated from Genetika, Vol. 41, No. 7, 2005, pp. 943–953.Original Russian Text Copyright © 2005 by Nikitina, Lebedev, Sukhanova, Nazarenko.     

Effects of Dietary Korean Proso-Millet Protein on Plasma Adiponectin,HDL Cholesterol,Insulin Levels,and Gene Expression in Obese Type 2 Diabetic Mice

We investigated the effect of dietary Korean proso-millet protein concentrate (PMP) on glycemic responses, plasma lipid levels, and the plasma level and gene expression of adiponectin in obese type 2 diabetic mice under normal and high-fat feeding conditions. The findings were that the feeding of PMP clearly elevated plasma high-density lipoprotein cholesterol (HDL cholesterol) and adiponectin levels and brought about effective reduction in the levels of glucose and insulin in mice under high-fat diet conditions as compared with a control diet. Gene expression study revealed that the diet up-regulated expression of adiponectin and down-regulated tumor necrosis factor-α (TNF-α). Considering the central role of adiponectin and HDL cholesterol in improving and ameliorating type 2 diabetes, obesity, and cardiovascular disease, our findings imply that PMP may have potential for therapeutic intervention in type 2 diabetes.     

Statistical analysis of variations in total cholesterol, HDL, HDL2, HDL3, LDL-cholesterol, VLDL-triglycerides, apo-A and apo-B lipoproteins in dyslipidemias during antilipemic therapy

In our study, bezafibrate in short-acting formula and long-acting formula, administered to hyperlipidaemic patients, resulted in a significant lowering of atherogenic lipids and lipoproteins (chol.-T, LDL and VLDL-chol., apolipoprotein B and VDLD-TR), and a marked increase in the levels of HDL, HDL2, HDL3-chol. and apolipoprotein A with a protective action as regards vascular atherosclerotic damage. The long-acting formula showed a greater effectiveness.     

Rare Mutations of CACNB2 Found in Autism Spectrum Disease-Affected Families Alter Calcium Channel Function

Autism Spectrum Disorders (ASD) are complex neurodevelopmental diseases clinically defined by dysfunction of social interaction. Dysregulation of cellular calcium homeostasis might be involved in ASD pathogenesis, and genes coding for the L-type calcium channel subunits Ca_V1.2 (CACNA1C) and Ca_Vβ2 (CACNB2) were recently identified as risk loci for psychiatric diseases. Here, we present three rare missense mutations of CACNB2 (G167S, S197F, and F240L) found in ASD-affected families, two of them described here for the first time (G167S and F240L). All these mutations affect highly conserved regions while being absent in a sample of ethnically matched controls. We suggest the mutations to be of physiological relevance since they modulate whole-cell Ba²⁺ currents through calcium channels when expressed in a recombinant system (HEK-293 cells). Two mutations displayed significantly decelerated time-dependent inactivation as well as increased sensitivity of voltage-dependent inactivation. In contrast, the third mutation (F240L) showed significantly accelerated time-dependent inactivation. By altering the kinetic parameters, the mutations are reminiscent of the CACNA1C mutation causing Timothy Syndrome, a Mendelian disease presenting with ASD. In conclusion, the results of our first-time biophysical characterization of these three rare CACNB2 missense mutations identified in ASD patients support the hypothesis that calcium channel dysfunction may contribute to autism.     

Evaluation of serum lipids and high-density lipoprotein subfractions (HDL2, HDL3) in postmenopausal patients with breast cancer

Breast cancer patients are known to be at increased risk for developing other chronic diseases including cardiovascular disease. Studies by different investigators have shown a correlation between increased dietary fat or hypercholesterolemia and the occurrence of breast cancer. Since previous studies on lipoprotein subfractions in this type of cancer have been inconsistent, we evaluated the lipids and lipoprotein subfraction levels in postmenopausal patients with breast cancer in an attempt to identify the risk for the development of cardiovascular disease.The study included 132 patients, 56 of which were suffering from breast cancer, 32 from pancreatic and 44 age-matched controls. Total cholesterol (TC), triglycerides and lipoprotein fractions as well as TC/High density lipoprotein (HDL) and HDL2/HDL3 ratios were estimated by standard laboratory techniques.An increase in triglycerides and a decrease in HDL-cholesterol, especially in the HDL2 subfraction, were observed in patients with breast cancer as compared to the controls (P < 0.05). The maximum changes in TC, and HDL concentrations were observed in patients with advanced disease. Analysis of indexes of atherosclerosis (TC/HDL, and HDL2/HDL3 ratios) demonstrated that breast cancer patients had significantly higher TC/HDL ratio (6.44 ± 1.24) compared with controls (3.43 ± 0.57, p = 0.001), and patients with pancreatic cancer (3.79 ± 0.15, p = 0.027).The results have demonstrated an unfavourable lipid profile in untreated breast cancer patients with high atherosclerosis indexes. This observation is of great importance, considering the potential use of endocrine therapy that could result in further deterioration of lipid indexes. We propose the evaluation and monitoring of lipid profile prior and after the induction of hormonal therapy in breast cancer patients, as a routine in clinical setting. (Mol Cell Biochem 268: 19–24, 2005)     

Association of RBP4 Gene Variants and Serum HDL Cholesterol Levels in the Newfoundland Population

Retinol‐binding protein 4 (RBP4) is a novel adipokine that likely contributes to systemic insulin resistance and dyslipidemia. The role of genetic variations in RBP4 on phenotypes of glucose and lipid metabolism is not clear in humans. The purpose of this study was to examine five single‐nucleotide polymorphisms (SNPs) in the RBP4 gene to determine their relationship with markers of insulin resistance and serum lipids in the CODING Study. The CODING Study consists of 1,836 subjects recruited from the genetically homogeneous population of Newfoundland and Labrador (NL), Canada. Serum glucose, insulin, homeostasis model assessment of insulin resistance (HOMA_IR), HOMA for β cell function (HOMA_β), total cholesterol (Chol), high‐density lipoprotein cholesterol (HDL‐C), low‐density lipoprotein cholesterol (LDL‐C), and triglycerides were determined after a 12‐h fast. Five SNPs within RBP4 (rs3758539, G/A 5′ flanking region; rs61461737, A/G intron; rs10882280, C/A intron; rs11187545, A/G intron; and rs12265684, C/G intron) were genotyped using TaqMan validated or functionally tested SNP genotyping assays. After correcting for multiple testing, we observed a significant association between the minor allele of two noncoding SNPs (rs10882280 and rs11187545) and higher serum HDL‐C (P = 0.043 and 0.042, respectively). No significant associations were observed with any other parameter related to lipid metabolism. We also found no significant association between any variant sites and markers of insulin resistance. Our results suggest that genetic variations in RBP4 may play a role in the differences in serum HDL‐C levels in the NL population.     

Effects of Methionine,Cystine and Taurine on Plasma Cholesterol Level in Rats Fed a High Cholesterol Diet

l-Methionine γ-lyase (EC 4.4.1.11) catalyzes α,β-elimination of l-2-amino-3-(N-methylamino)propionic acid and l-2-amino-3-(N-hydroxyethylamino)propionic acid to yield pyruvate, ammonia, and the corresponding amines. These amino acids also undergo the enzymatic β-replacement reaction with thiols to produce the corresponding S-substituted cysteines. Thus, l-methionine γ-lyase cleaves a C-N bond in addition to C-S, C-Se, and C-O bonds at the β position of amino acids by elimination and replacement reactions. A linear relationship between the reactivity, (log(V_max/Km) and the pKa value of the conjugated acid of the leaving group has been found for Se-methyl-l-selenocysteine, S-methyl-l-cysteine, and O-methyl-l-serine. However, l-2-amino-3-(N-methylamino)propionic acid has shown lower reactivity than that expected from the pKa value of methylammonium ions.     

Cholesterol synthesis in cultured human peripheral lymphocytes. Influence of LDL, HDL, cholesterol/phosphatidylcholine liposomes and complete serum

Lipoprotein-deficient milieu, freshly isolated human peripheral blood lymphocytes lose about 50% of their membrane cholesterol into the medium within 8 h. The cholesterol loss is counter-regulated by de novo synthesis commencing after a lag phase of 8-12 h, and reaching a steady state within 24 h at a diminished membrane cholesterol level. About 50 micrograms free cholesterol/ml, offered in the form of low-density lipoproteins (LDL) and cholesterol/phosphatidylcholine liposomes, suppressed cholesterol synthesis to about 20% of that controls (lipoprotein-deficient culture). By contrast, pure phosphatidylcholine liposomes enhanced cholesterol synthesis to about 150% of control values. High-density lipoproteins (HDL) exerted a slightly suppressive effect on cholesterol synthesis only at high concentrations (greater than 100 micrograms HDL cholesterol/ml). HDL added to cultures containing fixed concentrations of LDL led to a dose-dependent neutralization of LDL suppression of cholesterol synthesis. Culture medium containing complete serum caused a suppression of cholesterol synthesis to about 50% of the control. The lesser reduction in cholesterol synthesis caused by complete serum compared with LDL or cholesterol/phosphatidylcholine liposomes can be explained by the presence of HDL in the former. Our results support the view that the cholesterol requirement of blood lymphocytes in their lipid-rich milieu is met by cholesterol neosynthesis as well as an exchange mechanism with surrounding lipoproteins. In our system, the cholesterol neosynthesis appears to be controlled by the ratio of LDL to HDL in the surrounding medium.