Correlation of fasting serum apolipoprotein B-48 with coronary artery disease prevalence.

BACKGROUND: Postprandial hyperlipidemia partially refers to the postprandial accumulation of chylomicrons and chylomicron remnants (CM-R). Many in vitro studies have shown that CM-R has highly atherogenic properties, but consensus is lacking on whether CM-R accumulation correlates with the development of atherosclerotic cardiovascular diseases. We investigated the correlation between CM-R accumulation and the prevalence of coronary artery disease (CAD). DESIGN: Subjects who received a coronary angiography and did not take any lipid-lowering drugs (n = 189) were enrolled. Subjects with coronary artery stenosis (≥ 75%) were diagnosed as CAD. Biochemical markers for glucose and lipid metabolism including fasting apolipoprotein (apo) B-48 concentration were compared between CAD patients (n = 96) and age-, sex-, and body mass index (BMI)-matched non-CAD subjects without overt coronary stenosis (< 75%) (n = 67). We tried to determine which metabolic parameters were correlated with the prevalence of CAD by multiple logistic regression analysis, and whether or not the combination of high apo B-48 and other coronary risk factors (high triglyceride, low HDL-C, high HbA1c or low adiponectin levels) increased the prevalence of CAD. RESULTS: Fasting serum apo B-48 levels were significantly higher in CAD patients than in non-CAD subjects (3·9 ± 2·4 vs. 6·9 ± 2·6 µg/mL, P < 0·0001) and had the most significant correlation with the existence of CAD. The clustering of high fasting apo B-48 levels (> 4·34 µg/mL, the cut-off value) and other coronary risk factors were found to be associated with a stronger risk of CAD compared with single high fasting apo B-48 levels. CONCLUSION: Fasting serum apo B-48 levels significantly correlated with the prevalence of CAD.

Prognostic significance of the hair follicle stem cell marker nestin in patients with malignant melanoma.

Nestin is an intermediate filament protein, and serves as a hair follicle stem cell and neural stem cell marker. Recent studies have suggested that nestin expression is also important for tumorigenesis. Previous reports from our laboratory have revealed that nestin is a marker of HMB-45-negative melanoma cells in dermal invasive lesions of nodular malignant melanoma. The present study examines nestin expression in malignant melanoma and investigates the relationship between nestin expression and prognosis in patients. We immunohistochemically stained 78 formalin-fixed and paraffin-embedded malignant melanomas for nestin, HMB-45 and S100 reactivity. We found that nestin, HMB-45 and S100 protein were detected in 56.5%, 88.4% and 100% of malignant melanomas, respectively. The 5-year survival rate of stage I and II nestin-positive cases was significantly decreased compared to the nestin-negative cases (p < 0.05). In addition, the 5-year survival rate exceeded 80% in nestin-negative malignant melanomas at all stages of tumor development. We conclude that nestin expression may be a predictor of poor prognosis in patients with malignant melanoma.

Angiopoietin-like protein3 regulates plasma HDL cholesterol through suppression of endothelial lipase.

OBJECTIVE: A low level of high-density lipoprotein (HDL) in plasma has been recognized as an aspect of metabolic syndrome and as a crucial risk factor of cardiovascular events. However, the physiological regulation of plasma HDL levels has not been completely defined. Current studies aim to reveal the contribution of angiopoietin-like protein3 (angptl3), previously known as a plasma suppressor of lipoprotein lipase, to HDL metabolism. METHODS AND RESULTS: Angptl3-deficient mice showed low plasma HDL cholesterol and HDL phospholipid (PL), and which were increased by ANGPTL3 supplementation via adenovirus. In vitro, ANGPTL3 inhibited the phospholipase activity of endothelial lipase (EL), which hydrolyzes HDL-PL and hence decreases plasma HDL levels, through a putative heparin-binding site in the N-terminal domain of ANGPTL3. Post-heparin plasma in Angptl3-knockout mice had higher phospholipase activity than did that in wild-type mice, suggesting that the activity of endogenous EL is elevated in Angptl3-deficient mice. Furthermore, we established an ELISA system for human ANGPTL3 and found that plasma ANGPTL3 levels significantly correlated with plasma HDL cholesterol and HDL-PL levels in human subjects. CONCLUSIONS: Angptl3 acts as an inhibitor of EL and may be involved in the regulation of plasma HDL cholesterol and HDL-PL levels in humans and rodents.

Novel mutations of the HOXD13 gene in hand and foot malformations.

Homeobox genes encode a set of transcription factors of fundamental importance for body patterning during embryogenesis. Hoxa9-a13 and Hoxd9-d13 play an especially important part in vertebrate limb development. Synpolydactyly (SPD) is characterized by various malformations of the limbs. The expansion of the polyalanine tract in 1OXD13 is one of its major causes. Recently, there have been many analysis studies of HOXD13 in patients with SPD and limb malformations. We analyzed HOXD13 in 100 patients with limb malformations, which affects the limbs in the distal parts of the metacarpal and/or metatarsal bones. Seven mutations in the coding region and two mutations in the 5'-untranslated region were identified. All were novel mutations. In this study, the mutations were located upstream in the homeobox. Thus, translation of the homeobox was affected by upstream mutations. Consequently, this suggested the possibility that abnormalities in the hands and feet could be caused by novel HOXD13 gene mutations.

Identification of unique lipoprotein subclasses for visceral obesity by component analysis of cholesterol profile in high-performance liquid chromatography.

OBJECTIVE: The contribution of visceral fat accumulation to the development of coronary heart disease was previously reported, but the relation between visceral fat accumulation and serum lipoprotein subclasses was unknown. METHODS AND RESULTS: We examined the relation of lipoprotein subclasses with visceral fat accumulation in 62 male subjects (aged 22 to 67 years) with visceral fat syndrome or obesity. Cholesterol levels in very low-density, low-density, and high-density lipoprotein subclasses (VLDL, LDL, and HDL) were determined by computer-assisted high-performance liquid chromatography. Subcutaneous fat area and visceral fat area were measured by computed tomographic scanning. There was no significant correlation between the subcutaneous fat area and the cholesterol levels in all lipoprotein subclasses. In contrast, the visceral fat area was correlated positively (P<0.002) with VLDL and LDL subclasses, except for large LDL, but negatively (P<0.001) with those in large and medium HDL subclasses. The observed positive correlations of small and very small LDL subclasses remained significant (P<0.005) after adjustment for serum cholesterol, triglycerides, HDL cholesterol, and LDL cholesterol, respectively, but a significant negative correlation (P<0.005) of large LDL was obtained after adjustment for LDL cholesterol. CONCLUSIONS: These findings indicate that this simple high-performance liquid chromatography method may be applied for easy detection and evaluation of abnormal distribution of lipoprotein subclasses.

RECS1 deficiency in mice induces susceptibility to cystic medial degeneration.

RECS1 is a novel shear stress-responsive gene that encodes a protein putatively forming seven-span transmembrane domains. We reports here that mouse RECS1 (mRECS1) transcripts is detected in most tissues except for thymus, spleen and testis. The putative cytoplasmic N-terminus of mRECS1 has a high content of proline (23%) and glycine (12%) residues, contains one PPXY motif, multiple PXXP motifs and one overlapping P(T/S)AP and PPXY motif (P(T/S)APPXY). The PPXY motif lies within one potential PEST sequence (PEST score: +7.65). We prepared anti-RECS1 polyclonal antibody and found by western blot analysis that the mRECS1 protein in the lung and aorta was detected as a 34.4 kDa band. However, one shifted 58 kDa band or three shifted bands (48, 69, 82 kDa) were detected in the heart or the liver, respectively. Since northern blot detected only one species of mRECS1 mRNA in heart and liver tissues, as well as other tissues (approximately 2.2 kb), these differences in molecular weight seem to be due to posttranslational modification. Biochemical fractionation and RECS1-GFP fusion protein revealed that RECS1 localizes at the endosomal/lysosomal membranes in the cytoplasm. To understand the function of RECS1 in the body, we made RECS1 knockout (KO) mice and found that RECS1 KO mice (older than 14 months) are prone to cystic medial degeneration (CMD). Taken together, we conclude that RECS1 is an endosomal/lysosomal membrane protein which plays protective roles in vascular remodeling.

Matrix metalloproteinases as novel disease markers in Takayasu arteritis.

BACKGROUND: Takayasu arteritis (TA) is a chronic vasculitis that primarily affects large elastic arteries. Monitoring of disease activity is crucial because the disease tends to progress despite treatment with glucocorticoid and/or immunosuppressive agents. Erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) have generally been used as disease activity markers, but these are nonspecific inflammatory markers and lack the sensitivity and specificity to accurately monitor the disease status. Given the histological findings characterized by destruction of elastic fibers, we hypothesized that matrix metalloproteinases (MMPs) could be useful as markers of disease activity in TA. METHODS AND RESULTS: A consecutive series of 25 patients with TA were enrolled in this study. According to the National Institutes of Health criteria of disease activity, 11 were in an active phase and the remaining 14 were in remission. Circulating levels of MMP-2, MMP-3, and MMP-9 were determined by ELISA in all patients with TA and controls. MMP-2 levels were higher in patients with TA than in controls, but no correlation was found between serum MMP-2 and disease activity score. In contrast, MMP-3 and MMP-9 levels in patients with active disease were higher than in patients in remission and controls, and a positive correlation was demonstrated between circulating levels of MMP-3 or MMP-9 and disease activity score. The high levels of MMP-3 and MMP-9 improved when patients underwent remission. CONCLUSIONS: The present results indicate that MMP-2 can be helpful in diagnosing TA and that MMP-3 and MMP-9 can be used as activity markers for TA.

Adiponectin reduces atherosclerosis in apolipoprotein E-deficient mice.

BACKGROUND: Dysregulation of adipocyte-derived bioactive molecules plays an important role in the development of atherosclerosis. We previously reported that adiponectin, an adipocyte-specific plasma protein, accumulated in the injured artery from the plasma and suppressed endothelial inflammatory response and vascular smooth muscle cell proliferation, as well as macrophage-to-foam cell transformation in vitro. The current study investigated whether the increased plasma adiponectin could actually reduce atherosclerosis in vivo. METHODS AND RESULTS: Apolipoprotein E-deficient mice were treated with recombinant adenovirus expressing human adiponectin (Ad-APN) or beta-galactosidase (Ad-betagal). The plasma adiponectin levels in Ad-APN-treated mice increased 48 times as much as those in Ad-betagal treated mice. On the 14th day after injection, the lesion formation in aortic sinus was inhibited in Ad-APN-treated mice by 30% compared with Ad-betagal-treated mice (P<0.05). In the lesions of Ad-APN-treated mice, the lipid droplets became smaller compared with Ad-betagal-treated mice (P<0.01). Immunohistochemical analyses demonstrated that the adenovirus-mediated adiponectin migrate to foam cells in the fatty streak lesions. The real-time quantitative polymerase chain reaction revealed that Ad-APN treatment significantly suppressed the mRNA levels of vascular cell adhesion molecule-1 by 29% and class A scavenger receptor by 34%, and tended to reduce levels of tumor necrosis factor-alpha without affecting those of CD36 in the aortic tissue. CONCLUSIONS: These findings documented for the first time that elevated plasma adiponectin suppresses the development of atherosclerosis in vivo.

Human scavenger receptor class B type I is expressed with cell-specific fashion in both initial and terminal site of reverse cholesterol transport.

The reverse cholesterol transport (RCT) is one of the major protective systems against atherosclerosis, in which high-density lipoprotein (HDL) removes cholesterol from lipid-laden cells and delivers it to the liver. Scavenger receptor class B type I (SR-BI) is a HDL receptor in the liver and adrenal glands and is involved in the selective uptake of cholesteryl ester from HDL, which has been extensively, analyzed using rodent models. However, the expression and regulation of the human homologue of this receptor are not known yet. We previously reported that this receptor is expressed in in vitro differentiated macrophages and its expression is up-regulated by the addition of modified lipoproteins into the medium [Hirano K, Yamashita S, Nakagawa Y, et al. Expression of human scavenger receptor class B type I in cultured human monocyte-derived macrophages and atherosclerotic lesions. Circ Res 1999;85:108-16]. In order to further investigate the physiological significance of this receptor in humans, we have performed extensive immunohistochemical analyses with specimens of the liver and adrenal glands as well as arteries with different stages of atherosclerotic lesions. In human liver and adrenal glands, a positive SR-BI immunoreactivity was detected in both hepatic and adrenal parenchymal cells as well as Kupffer cells. These parenchymal cells had a strong signal on the cell surface, whereas Kupffer cells showed a heterogeneous and punctate pattern. In human aorta and coronary arteries, SR-BI was highly expressed in atherosclerotic plaques, but not in non-atherosclerotic lesions. Double immunostaining revealed that SR-BI was expressed in a subpopulation of macrophages, of which staining pattern was similar to that observed in Kupffer cells. These data clearly demonstrated that SR-BI was expressed with cell-specific fashions in both the initial and terminal step of RCT in humans. Thus, SR-BI might be physiologically relevant and have distinct tissue-specific functions.

Defective cholesterol efflux in Werner syndrome fibroblasts and its phenotypic correction by Cdc42, a RhoGTPase.

Werner syndrome (WS) is characterized by the early onset of senescent phenotypes including premature atherosclerotic cardiovascular diseases, although the underlying molecular mechanism for atherosclerosis has not been fully understood yet. Cholesterol efflux from the cells is the initial step of reverse cholesterol transport, a major protective system against atherosclerosis. The aim of the present study was to determine whether this crucial step may be altered in WS. We examined intracellular lipid transport and cholesterol efflux and the expression levels of its related molecules in skin fibroblasts obtained from patients with WS. Cholesterol efflux was markedly reduced in the WS fibroblasts in association with increased cellular cholesterol. Fluorescent recovery after photobleaching (FRAP) technique revealed that intracellular lipid transport around Golgi apparatus was markedly reduced when using a C6-NBD-Ceramide as a tracer. Cdc42 protein and its GTP-bound form were markedly reduced in the WS fibroblasts. The complementation of wild-type Cdc42 corrected cholesterol efflux, intracellular lipid transport, and cellular cholesterol levels in the WS fibroblasts. These data indicated that the reduced expression of Cdc42 may be responsible for the abnormal lipid transport, which in turn might be related to the cardiovascular manifestations in WS.

Familial massive tendon xanthomatosis with decreased high-density lipoprotein-mediated cholesterol efflux.

We experienced a family with novel massive tendon xanthomatosis which can be excluded from known disease causing xanthomatosis. The proband was a 58-year-old man who had necrosis in his massive Achilles tendon xanthoma. Three of 5 brothers including him and his nephew had the same clinical phenotype. The systemic tendon xanthomatosis became apparent around 30 years of their age. The proband and his elder brother had mild elevations of serum total cholesterol level (251 and 228 mg/dL, respectively). The low-density lipoprotein receptor activity of the proband's lymphocytes was normal. Neither plant sterol nor cholestanol level was increased in the proband's plasma. Magnetic resonance image of the proband's Achilles tendon demonstrated a massive expansion of the soft tissue with salami sausage-like appearance in his heels (50 mm in thickness). The physiological function of macrophages (MPhi) from the patients was investigated to clarify the mechanism for the formation of xanthomatosis. There was no significant difference in the uptake of oxidized low-density lipoprotein between the proband's MPhi and the control. High-density lipoprotein 3-mediated cholesterol efflux from the patients' MPhi (n = 2) was significantly reduced compared with the controls (n = 3), whereas there was no difference in apolipoprotein (apo) A-I-mediated cholesterol efflux between the patients' MPhi and the controls. Furthermore, there was no reduction of the messenger RNA levels of ATP-binding cassette transporter 1 (ABCA1), which is involved in apo A-I-mediated cholesterol efflux, in the proband's MPhi compared with the control. The present study demonstrates that the mechanism for the formation of novel familial massive tendon xanthomatosis may be, at least in part, associated with decreased high-density lipoprotein 3, but not free apo A-I-mediated cholesterol efflux from MPhi in vivo.

Carriers of the frequent lipoprotein lipase S447X variant exhibit enhanced postprandial apoprotein B-48 clearance.

The frequent lipoprotein lipase S447X variant (LPLS447X) is firmly associated with a lower incidence of cardiovascular disease, the mechanisms for which remain to be established. To further unravel these beneficial effects, we studied the consequences of LPLS447X heterozygosity on LPL mass and activity, as well as on the postprandial lipoprotein profile. Fifteen male heterozygous LPLS447X carriers and 15 matched control subjects received an oral fat load (30 g/m(2)). Lipid parameters were evaluated at baseline and 2, 3, 4, and 6 hours after fat loading. LPL concentration and activity were analyzed, and endothelial function was evaluated noninvasively as flow-mediated dilation of the brachial artery. Although baseline apoprotein B-48 (apoB48) levels were similar, the rise in apoB48 was attenuated in LPLS447X carriers with 25% lower peak values compared with controls (P=.04). In conjunction, LPLS447X carriers were characterized by a 2.4-fold increase in pre-heparin LPL mass (P<.0001). The decrease in postprandial flow-mediated dilation was comparable in both groups. LPLS447X carriers exhibit enhanced apoB48 clearance with concomitant increase in pre-heparin LPL mass, without changes in LPL activity. This combination might suggest a role for increased ligand action of LPL in LPLS447X carriers contributing to the cardiovascular protection in carriers of this mutation.

Apolipoprotein B-100 kinetics and static plasma indices of triglyceride-rich lipoprotein metabolism in overweight men.

OBJECTIVE: We examined the association of plasma apolipoprotein (apoB) B-48, remnant-like particle (RLP)-cholesterol and non-HDL cholesterol concentrations with apoB-100 kinetics in overweight-obese men. METHODS AND RESULTS: Very-low density lipoprotein (VLDL), intermediate-density lipoprotein (IDL) and low-density lipoprotein (LDL) apoB-100 kinetics were measured in 53 men (BMI 33 +/- 4 kg/m(2)) using stable isotopes and multicompartmental modeling to estimate production rate (PR) and fractional catabolic rate (FCR). Fasting apoB-48 and RLP-cholesterol concentrations were measured using immunoassays. In univariate regression, apoB-48 and RLP-cholesterol were inversely associated with VLDL-apoB-100 FCR and IDL-apoB-100 FCR (P < 0.01 for all), but not with VLDL-, IDL- and LDL-apoB-100 PRs. Plasma non-HDL-cholesterol concentration was significantly and positively associated with the secretion rate of VLDL-apoB-100 (P < 0.05), and inversely correlated with the FCR of LDL-apoB-100 (P < 0.01). CONCLUSIONS: Our findings suggest that in overweight-obese men plasma concentrations of apoB-48, RLP-cholesterol and non-HDL-cholesterol are partly dependent on catabolism of apoB-100 containing lipoproteins.

Retarded intracellular lipid transport associated with reduced expression of Cdc42, a member of Rho-GTPases, in human aged skin fibroblasts: a possible function of Cdc42 in mediating intracellular lipid transport.

OBJECTIVE: Many cell types in atherosclerotic lesions are thought to have various biological abnormalities, such as impaired lipid homeostasis and slow cell proliferation, which may be related to senescence at cellular and individual levels. One of the common characteristics of senescent cells in vitro is the alteration of actin cytoskeletons, which have been reported to be involved in the intracellular transport of lipids. Recently, we raised the hypothesis that Cdc42, which is a member of the Rho-GTPase family and is known to play an important role in actin dynamics, might be important in cellular lipid transport. METHODS AND RESULTS: In the present study, we found that the protein expression levels and GTP-binding activities of Cdc42 were decreased in aged human skin fibroblasts. Moreover, we found the intracellular kinetics of Golgi-associated lipids to be retarded in these cells, which was demonstrated by the fluorescence recovery after photobleaching (FRAP) technique and the use of N-[7-(4-nitrobenzo-2-oxa-1,3-diazole)]-6-aminohexanoyl-D-erythro-sphingosine as a tracer. To correlate the decreased expression of Cdc42 with the retarded FRAP, we complemented the amount of wild-type c-myc-tagged Cdc42Hs (myc-Cdc42Hs-WT) by adenovirus-mediated gene transfer. We further tested the effect of the dominant-active form (myc-Cdc42Hs-DA, V12Cdc42Hs) or dominant-negative form (myc-Cdc42Hs-DN, N17Cdc42Hs) of Cdc42Hs on FRAP. Introduction of myc-Cdc42Hs-WT or myc-Cdc42Hs-DA recovered the retarded FRAP in the aged fibroblasts. Conversely, control fibroblasts infected with myc-Cdc42Hs-DN exhibited significantly retarded FRAP. CONCLUSIONS: These data clearly indicate that the expression of Cdc42, a small G protein, is decreased in the aged cells in close association with the retarded intracellular lipid transport. The present study demonstrates a possible function of Cdc42 in the mediation of intracellular lipid transport.

Atorvastatin markedly improves type III hyperlipoproteinemia in association with reduction of both exogenous and endogenous apolipoprotein B-containing lipoproteins.

Remnant lipoproteins are known to promote atherosclerosis especially in patients with type III hyperlipoproteinemia (HLP). In the current study, the effects of atorvastatin were investigated with special reference to the exogenous and endogenous apolipoprotein (apo) B-containing lipoprotein metabolism in type III HLP. Four Japanese male patients with type III HLP associated with homozygous apoE2 were studied. One-month administration of atorvastatin (20 mg once daily), after a 4-week dietary run-in, strikingly reduced serum total cholesterol and triglyceride (TG) levels by 52 (P<0.01) and 56% (P<0.05), respectively. Atorvastatin further decreased remnant-like particle (RLP)-cholesterol by 73% and RLP-TG by 65% (P<0.05), respectively. Distribution analysis by polyacrylamide gel disc electrophoresis clearly showed that atorvastatin diminished very low-, intermediate- and low-density lipoprotein particles. The relative particle diameter of intermediate-density lipoprotein became smaller after atorvastatin treatment (P<0.01). Furthermore, ultracentrifugal analysis demonstrated that atorvastatin significantly decreased cholesterol, TG and phospholipid concentrations in all apoB-containing lipoprotein fractions and very low-density lipoprotein (VLDL)-cholesterol/serum TG ratio (P<0.05), implying atorvastatin-induced reduction of beta-VLDL. Finally, newly developed assays of apoB-48 and apoB-100 revealed that atorvastatin markedly reduced these apolipoproteins by 43 and 52%, respectively (P<0.01), suggesting that atorvastatin decreased the number of both exogenous and endogenous apoB-containing lipoproteins. Taken together, atorvastatin improves remnant lipoprotein metabolism in type III HLP both in quality and in quantity. Atorvastatin can be one of the optimal options for the treatment of patients with type III HLP.

Large scale isolation of non-uniform shear stress-responsive genes from cultured human endothelial cells through the preparation of a subtracted cDNA library.

To investigate the molecular mechanisms responsible for the regional selectivity of early atherogenesis, we have applied a non-uniform shear stress to cultured human umbilical vein endothelial cells (HUVEC). We used a microcarrier culture system and a combination of subtraction and reverse-subtraction methods to isolate a number of genes upregulated by shear stress. The resultant subtracted library includes several known genes (e.g. MCP-1, TM) whose responsiveness to shear stress has been previously reported, indicating that the library is enriched for genes upregulated by shear stress. Also included are atherosclerosis-related genes (e.g. CTGF, IL-8) whose responsiveness to shear stress had not been demonstrated, other known genes whose relationship to atherosclerosis had not been reported, and novel genes. Some responsive to centrifugal force and shear stress (RECS) genes are also upregulated following stimulation by steady laminar shear stress in a parallel plate chamber. Interestingly, the library includes ET-1 and PAI, which are well known atherogenic factors that are downregulated by laminar shear stress. This implies that turbulent shear stress has effects on HUVEC that are different from those elicited by laminar shear stress. Importantly, analysis of specimens taken from human aorta showed that several RECS genes are transcriptionally upregulated in atherosclerotic lesions, suggesting that the subtracted library includes novel therapeutic targets for the treatment of atherosclerosis.

Prevalence and phenotypic spectrum of cholesteryl ester transfer protein gene mutations in Japanese hyperalphalipoproteinemia.

A patient with cholesteryl ester transfer protein (CETP) deficiency presents with marked hyperalphalipoproteinemia (HALP). To investigate the contribution of CETP deficiency to the cause of HALP (HDL-C> or =1.94 mmol/l, 75 mg/dl), we investigated the CETP activities and the prevalence of genetic CETP mutations among 624 Japanese HALP subjects. The subjects were screened for four known genetic CETP mutations (intron 14 splicing defect (In14), exon 15 missense mutation (Ex15), intron 10 splicing defect (In10) and exon 6 nonsense mutation (Ex6)). We found the frequency of the patients with reduced CETP activity (<75% of normal controls) to be 55.5 and 64.1% in a high HDL group (1.94< or =HDL-C<2.59 mmol/l) and a marked HALP group (HDL-C> or =2.59 mmol/l, 100 mg/dl), respectively. At least one of the four mutations was identified in 65.7% of subjects with reduced CETP activities and 57.5% of subjects with marked HALP. The In14 and Ex15 mutations were very common in HALP subjects and the frequency of In10 mutation and Ex6 mutation was quite low. To investigate the impact of genetic CETP mutation on the phenotypes, we compared the plasma lipid levels and CETP activities between the subjects with two common mutations. All In14 homozygotes showed marked HALP, while marked HALP is less frequent (64.3%) in Ex15 homozygotes. HDL-C levels in Ex15 heterozygotes were significantly higher than those of In14 heterozygotes, suggesting the mutation has dominant negative effects on CETP activity in vivo. Some cases with In14 (5.7%) or Ex15 (7.2%) mutation showed low HDL-C levels. We conclude that CETP deficiency is a major cause of HALP; nevertheless CETP deficiency is not necessarily HALP.

Molecular mechanisms of cholesteryl ester transfer protein deficiency in Japanese.

Plasma cholesteryl ester transfer protein (CETP) facilitates the transfer of cholesteryl ester (CE) from high density lipoprotein (HDL) to apolipoprotein B-containing lipoproteins. Since CETP regulates the plasma levels of HDL cholesterol and the size of HDL particles, CETP is considered to be a key protein in reverse cholesterol transport (RCT), a protective system against atherosclerosis. The importance of plasma CETP in lipoprotein metabolism was demonstrated by the discovery of CETP-deficient subjects with marked hyperalphalipoproteinemia (HALP). Genetic CETP deficiency is the most important and common cause of HALP in the Japanese. Ten mutations of the CETP gene have been demonstrated as causes of HALP, including two common mutations: an intron 14 splicing defect (Int14 + 1 G --> A) and an exon 15 missense mutation (D442G). The subjects with CETP deficiency show a variety of abnormalities in the concentration, composition, and function of both HDL and low density lipoprotein (LDL). CETP deficiency is considered a physiological state of impaired RCT, which may possibly lead to the development of atherosclerosis despite high HDL cholesterol levels. However, the pathophysiological significance of CETP in terms of atherosclerosis has been controversial. Epidemiological studies in Japanese-Americans living in Hawaii and Japanese in the Omagari area, where HALP subjects with an intron 14 splicing defect of the CETP gene are markedly frequent, have shown a relatively increased incidence of coronary atherosclerosis in CETP deficiency. On the other hand, the TaqIB polymorphism-B2 allele with low CETP mass and increased HDL cholesterol has been related to a decreased risk for coronary heart disease (CHD) in many studies, including the Framingham Offspring Study. The current review focused on the characterization of the Japanese subjects with CETP deficiency, including our recent findings.