Acute encephalitis with refractory, repetitive partial seizures (AERRPS): a peculiar form of childhood encephalitis.

OBJECTIVE: We conducted a nationwide multicenter study in Japan to elucidate the clinical and laboratory characteristics of acute encephalitis with refractory, repetitive partial seizures (AERRPS). MATERIALS AND METHODS: Clinical and laboratory features, treatment, and outcome were assessed using a structured questionnaire. RESULTS: Twenty-nine children were enrolled in the study. Refractory and repetitive partial seizures accompanied by fever were the cardinal clinical features. Partial seizures consisted principally of eye deviation or facial twitching, being periodically repeated during the acute phase. These seizures were refractory to conventional anticonvulsants and were only suppressed by high-dose intravenous barbiturate administration. Rhythmic activities on electroencephalography and non-specific cerebral atrophy on neuroimaging were common. Serum or cerebrospinal antibodies against GluRepsilon2 were positive in six patients. General prognosis was unfavorable due to intractable epilepsy and cognitive deficits. CONCLUSION: Based on the peculiar and homogenous features, AERRPS can be regarded as a distinct clinical entity.

Lapaquistat acetate, a squalene synthase inhibitor, changes macrophage/lipid-rich coronary plaques of hypercholesterolaemic rabbits into fibrous lesions.

BACKGROUND AND PURPOSE: Inhibition of squalene synthesis could transform unstable, macrophage/lipid-rich coronary plaques into stable, fibromuscular plaques. We have here treated WHHLMI rabbits, a model for coronary atherosclerosis and myocardial infarction, with a novel squalene synthase inhibitor, lapaquistat acetate (TAK-475). EXPERIMENTAL APPROACH: Young male WHHLMI rabbits were fed a diet supplemented with lapaquistat acetate (100 or 200 mg per kg body weight per day) for 32 weeks. Serum lipid levels were monitored every 4 weeks. After the treatment, lipoprotein lipid and coenzyme Q10 levels were assayed, and coronary atherosclerosis and xanthomas were examined histopathologically or immunohistochemically. From histopathological and immunohistochemical sections, the composition of the plaque was analysed quantitatively with computer-assisted image analysis. Xanthoma was evaluated grossly. KEY RESULTS: Lapaquistat acetate decreased plasma cholesterol and triglyceride levels, by lowering lipoproteins containing apoB100. Development of atherosclerosis and xanthomatosis was suppressed. Accumulation of oxidized lipoproteins, macrophages and extracellular lipid was decreased in coronary plaques of treated animals. Treatment with lapaquistat acetate increased collagen concentration and transformed coronary plaques into fibromuscular plaques. Lapaquistat acetate also suppressed the expression of matrix metalloproteinase-1 and plasminogen activator inhibitor-1 in the plaque and increased peripheral coenzyme Q10 levels. Increased coenzyme Q10 levels and decreased very low-density lipoprotein cholesterol levels were correlated with improvement of coronary plaque composition. CONCLUSION AND IMPLICATIONS: Inhibition of squalene synthase by lapaquistat acetate delayed progression of coronary atherosclerosis and changed coronary atheromatous plaques from unstable, macrophage/lipid accumulation-rich, lesions to stable fibromuscular lesions.

Acute encephalopathy associated with influenza and other viral infections.

Acute encephalopathy is the most serious complication of pediatric viral infections, such as influenza and exanthem subitum. It occurs worldwide, but is most prevalent in East Asia, and every year several hundreds of Japanese children are affected by influenza-associated encephalopathy. Mortality has recently declined, but is still high. Many survivors are left with motor and intellectual disabilities, and some with epilepsy. This article reviews various syndromes of acute encephalopathy by classifying them into three major categories. The first group caused by metabolic derangement consists of various inherited metabolic disorders and the classical Reye syndrome. Salicylate is a risk factor of the latter condition. The second group, characterized by a systemic cytokine storm and vasogenic brain edema, includes Reye-like syndrome, hemorrhagic shock and encephalopathy syndrome, and acute necrotizing encephalopathy. Non-steroidal anti-inflammatory drugs, such as diclofenac sodium and mephenamic acid, may aggravate these syndromes. Severe cases are complicated by multiple organ failure and disseminated intravascular coagulation. Mortality is high, although methylprednisolone pulse therapy may be beneficial in some cases. The third group, characterized by localized edema of the cerebral cortex, has recently been termed acute encephalopathy with febrile convulsive status epilepticus, and includes hemiconvulsion-hemiplegia syndrome and acute infantile encephalopathy predominantly affecting the frontal lobes. Theophylline is a risk factor of these syndromes. The pathogenesis is yet to be clarified, but an increasing body of evidence points to excitotoxicity and delayed neuronal death.

Acute encephalopathy associated with influenza and other viral infections.

Acute encephalopathy is the most serious complication of pediatric viral infections, such as influenza and exanthem subitum. It occurs worldwide, but is most prevalent in East Asia, and every year several hundreds of Japanese children are affected by influenza-associated encephalopathy. Mortality has recently declined, but is still high. Many survivors are left with motor and intellectual disabilities, and some with epilepsy. This article reviews various syndromes of acute encephalopathy by classifying them into three major categories. The first group caused by metabolic derangement consists of various inherited metabolic disorders and the classical Reye syndrome. Salicylate is a risk factor of the latter condition. The second group, characterized by a systemic cytokine storm and vasogenic brain edema, includes Reye-like syndrome, hemorrhagic shock and encephalopathy syndrome, and acute necrotizing encephalopathy. Non-steroidal anti-inflammatory drugs, such as diclofenac sodium and mephenamic acid, may aggravate these syndromes. Severe cases are complicated by multiple organ failure and disseminated intravascular coagulation. Mortality is high, although methylprednisolone pulse therapy may be beneficial in some cases. The third group, characterized by localized edema of the cerebral cortex, has recently been termed acute encephalopathy with febrile convulsive status epilepticus, and includes hemiconvulsion-hemiplegia syndrome and acute infantile encephalopathy predominantly affecting the frontal lobes. Theophylline is a risk factor of these syndromes. The pathogenesis is yet to be clarified, but an increasing body of evidence points to excitotoxicity and delayed neuronal death.

Increase in hepatic low-density lipoprotein receptor activity during pregnancy in Watanabe heritable hyperlipidemic rabbits; an animal model for familial hypercholesterolemia.

In homozygous Watanabe heritable hyperlipidemic (WHHL) rabbits, the serum cholesterol level and serum low-density lipoprotein (LDL) level decreased from 562 +/- 76 (mean +/- S.E.) to 144 +/- 34 mg/dl and 410 +/- 56 to 90 +/- 25 mg/dl, respectively, during pregnancy, although the LDL receptor in this rabbit is genetically deficient. When Tyroxapol, which inhibits the degradation of very-low-density lipoprotein (VLDL), as well as Triton WR-1339, was injected into WHHL rabbits, the rate of the increase in serum cholesterol level in pregnant rabbits was not statistically different from that in non-pregnant rabbits. This result implied that the secretion rate of VLDL-cholesterol, the precursor of LDL-cholesterol, did not decrease during pregnancy. The amount of 125I-labeled LDL bound to LDL receptor was increased 1.8-fold in normal rabbits (from 29.3 +/- 4.3 to 52.3 +/- 4.6 ng/mg protein) and 12-fold in WHHL rabbits (from 0.5 +/- 0.2 to 6.0 +/- 0.7 ng/mg protein) during pregnancy. These results suggest that the decrease in serum cholesterol level in WHHL rabbits during pregnancy was associated with an increase in hepatic LDL receptor activity, which plays an important role in the regulation of serum cholesterol level.

High dose of fluvastatin sodium (XU62-320), a new inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, lowers plasma cholesterol levels in homozygous Watanabe-heritable hyperlipidemic rabbits.

The effects of fluvastatin sodium (XU62-320), a new type of inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, on plasma cholesterol and triacylglycerol levels were investigated using homozygous Watanabe-heritable hyperlipidemic (WHHL) rabbit, an LDL-receptor-deficient animal which expresses a hepatic LDL receptor activity less than 5% that of control rabbits. Plasma levels of total, VLDL- and LDL-cholesterol were decreased profoundly after oral administration of fluvastatin at a dose of 50 mg/kg per day for 4 weeks. Plasma triacylglycerol levels were not affected by fluvastatin. Hepatic HMG-CoA reductase activity increased by 3-fold and hepatic LDL receptor activity increased by only 3.7-fold, as calculated by Scatchard plot analysis, with fluvastatin administration for 4 weeks, and the hepatic mRNA level for the rabbit LDL receptor was increased by 3-fold. Combined administration of fluvastatin (50 mg/kg per day) and cholestyramine, a bile acid sequestrant resin, at a level of 2% of the diet for 4 weeks more profoundly decreased plasma total, VLDL- and LDL-cholesterol levels with induction of hepatic cholesterol 7 alpha-hydroxylase and no further induction of the hepatic LDL receptor. Plasma triacylglycerol levels were increased by the combination treatment. These results suggest that high dose of fluvastatin sodium is effective in lowering plasma cholesterol levels in homozygous WHHL rabbits through the shared mechanisms involving decrease in production and secretion of cholesterol from the liver and the induction of hepatic LDL receptor. Additional effect of cholestyramine on decrease in plasma cholesterol levels seems to be due to the further decrease in hepatic cholesterol secretion by up-regulation of hepatic cholesterol 7 alpha-hydroxylase.

Preventive effect of pravastatin sodium, a potent inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, on coronary atherosclerosis and xanthoma in WHHL rabbits.

In this paper, we examined whether the development of atherosclerosis in the Watanabe heritable hyperlipidemic (WHHL) rabbit, an animal model of familial hypercholesterolemia in man, could be prevented by the reduction of serum cholesterol levels. Pravastatin sodium (the generic name of CS-514), a potent inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, was used as a cholesterol-lowering drug. The drug was administered orally to 12 WHHL rabbits (2-3 months old) at a dose of 50 mg/kg per day for 24 weeks, and 13 animals were given water as control. In the treated group, serum cholesterol, phospholipid and triacylglycerol levels were significantly reduced by 28%, 32% and 16%, respectively, as compared with those of the control group. Although the prevention of development of the aortic atherosclerosis was not significant, the progression of coronary atherosclerosis was significantly prevented. The incidence of atherosclerosis in four main coronary arteries was reduced from 42% (control group) to 19% (treated group, P less than 0.01), and the development of lesion of coronary arteries evaluated by area of lesion was reduced from 19.7% (control group) to 9.1% (treated group, P less than 0.05). Histopathological findings supported the above observations. In addition, development of xanthoma in digital joints was also reduced from 90.4% (control group) to 58.3% (treated group, P less than 0.005). These results suggest that the development of coronary atherosclerosis and xanthoma in WHHL rabbit was reduced by continuous reduction of serum cholesterol levels treated with pravastatin sodium.

Statins alter smooth muscle cell accumulation and collagen content in established atheroma of watanabe heritable hyperlipidemic rabbits.

BACKGROUND: Acute coronary syndromes often result from rupture of vulnerable plaques. The collagen content of plaques probably regulates their stability. This study tested whether HMG-CoA reductase inhibitors (statins) alter interstitial collagen gene expression or matrix metalloproteinase (MMP) levels in rabbit atheroma. METHODS AND RESULTS: We administered equihypocholesterolemic doses of pravastatin (a hydrophilic statin, 50 mg. kg(-1). d(-1), n=9), fluvastatin (a cell-permeant lipophilic statin, 20 mg. kg(-1). d(-1), n=10), or placebo (n=10) to mature Watanabe heritable hyperlipidemic rabbits for 52 weeks. The fluvastatin group achieved a much higher peak plasma concentration (23.7 micromol/L) than did the pravastatin group (1.3 micromol/L) under these conditions. Immunohistochemistry revealed that MMP-1, MMP-3, and MMP-9 expression by macrophages in the intima was lower in both the pravastatin and fluvastatin groups than in the placebo group, whereas there was no difference in macrophage numbers. Numbers of intimal smooth muscle cells (SMCs) (identified by immunohistochemistry) and expression of type I procollagen mRNA (detected by in situ hybridization), however, were significantly higher in the pravastatin group than in the fluvastatin group. Treatment with pravastatin, but not fluvastatin, preserved interstitial collagen content in vivo (detected by picrosirius red polarization). In vitro, fluvastatin, but not pravastatin, decreased numbers of rabbit and human aortic SMCs without altering procollagen I mRNA expression. CONCLUSIONS: This study showed that statins can reduce MMP expression in atheroma and that cell-permeant statins can decrease SMC number and collagen gene expression in vivo.

Heme oxygenase-1 inhibits atherogenesis in Watanabe heritable hyperlipidemic rabbits.

BACKGROUND: Heme oxygenase-1 (HO-1) is proposed to have a variety of adaptive responses against oxidative stress. To examine the function of HO-1 against atherogenesis in vivo, we observed the effects of HO-1 inhibition on atherosclerotic lesion formation in Watanabe heritable hyperlipidemic rabbits (WHHL). Methods and Results- During 4 weeks of a 1% cholesterol diet, intravenous injections of Sn-protoporphyrin IX to inhibit HO-1 (S group, n=10) and saline as a control (C group, n=10) were given to 3-month-old WHHL rabbits. The percentages of en face atherosclerotic lesion areas in total descending aorta by Sudan IV staining (EFA) and the ratio of intima to media in microscopic atherosclerotic lesions in the ascending aortas (I/M) were calculated. Two different quantitative methods revealed significantly greater atherosclerotic lesions in the S group than the C group (EFA, P<0.001; I/M, P<0.005). HO-1 expression in atherosclerotic lesions was confirmed by Northern blot and immunohistochemical analyses. The dominant cell types expressing HO-1 were macrophages and foam cells, in which oxidized phospholipids were also accumulated. HO inhibition increased plasma and tissue lipid peroxide levels without affecting plasma lipid co osition. CONCLUSIONS: These results suggest the possibilities that HO-1 has antiatherogenic properties in vivo and that the antiatherogenic properties of HO-1 are conducted through the prevention of lipid peroxidation.

Granulocyte-macrophage colony-stimulating factor prevents the progression of atherosclerosis via changes in the cellular and extracellular composition of atherosclerotic lesions in watanabe heritable hyperlipidemic rabbits.

BACKGROUND: A cytokine network is involved in atherogenesis. This study was conducted to investigate the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) on the development and composition of atherosclerotic lesions in Watanabe heritable hyperlipidemic (WHHL) rabbits. METHODS AND RESULTS: GM-CSF (10 microg. kg-1. d-1) was administered to 4-month-old WHHL rabbits (n=9) 5 days a week for 7.5 months, whereas an equal dose of human serum albumin was administered to controls (n=9). The cholesterol levels were not changed significantly by the treatment. Age-matched 4-month-old rabbits (n=7) had atheromatous plaques over 30.7+/-5.7% of the inner surface area of the aortic arch. After treatment, the percentages of surface atheromatous plaques to total aortic arch area were 45.0+/-12.6% in the GM-CSF group and 74.3+/-11.0% in controls (P<0.0001). Histological examination demonstrated that GM-CSF reduced the ratio of intima to media (P<0.01) and cross-sectional areas of atherosclerotic lesions (P<0.0001). Quantitative analysis indicated a marked decrease in the areas of smooth muscle cells (P=0.0001), collagen (P=0.0001), and extracellular lipid deposits (P<0.05) of atheromatous plaques in GM-CSF-treated rabbits compared with controls. The terminal deoxynucleotidyltransferase-mediated dUTP-digoxigenin nick end-labeling (TUNEL) method and immunohistochemistry were performed to examine the relationship between decreased atherosclerotic lesions and apoptosis. The percentage of TUNEL-positive cells increased in the GM-CSF group (GM-CSF, 24.1+/-4.4% versus control, 11.6+/-3.2%; P<0.0001). GM-CSF enhanced the apoptosis of smooth muscle cells in the shoulder region and the fibrous cap (P<0.0001), suggesting one of the mechanisms for the antiatherogenic effect. CONCLUSIONS: GM-CSF altered the composition of atherosclerotic lesions and reduced the atherosclerosis in WHHL rabbits.

Serotonin-induced hypercontraction through 5-hydroxytryptamine 1B receptors in atherosclerotic rabbit coronary arteries.

BACKGROUND: Augmented vasoconstriction to serotonin (5-hydroxytryptamine [5-HT]) in atherosclerotic vessels plays a crucial role in the development of myocardial ischemia. We investigated mechanisms for serotonin-evoked hypercontraction in atherosclerotic rabbit coronary arteries. METHODS AND RESULTS: Contractile responses to serotonergic agents of endothelium-denuded coronary arteries from control and Watanabe heritable hyperlipidemic rabbits (WHHL) were examined. WHHL coronary arteries exhibited hypercontraction to 5-HT(1)-receptor agonists; the constrictor threshold concentrations and E:D(50) to serotonin, 5-carboxamidotryptamine, and sumatriptan in WHHL were significantly lower, and the E:(max) in WHHL to these agents were increased 55% to 59% above those of the control. Serotonin-evoked contractions in both groups were inhibited by GR127935 (5-HT(1B/1D) antagonist; 0.1 to 1 nmol/L) and pertussis toxin but not by ketanserin (5-HT(2) antagonist; 0.01 to 1 micromol/L), suggesting that the hypercontraction is most likely mediated by 5-HT(1B/1D) receptors through a pertussis toxin-sensitive pathway. Furthermore, simultaneous measurements of [Ca(2+)](i) and isometric tension of fura-2-loaded arteries revealed that the hypercontraction was concomitant with the augmented elevation of [Ca(2+)](i) in the smooth muscle. The 5-HT(1B) mRNA levels in WHHL coronary arteries increased to 2.5-fold over those in control arteries, whereas neither 5-HT(1D) nor 5-HT(2A) mRNA was detected in either group. CONCLUSIONS: Atherosclerotic rabbit coronary arteries exhibited the enhancement in contraction and Ca(2+) mobilization in response to serotonin. The 5-HT(1B) receptor, which is upregulated by atherosclerosis, most likely mediates the augmenting effects of serotonin.

MRI study of atherosclerotic plaque progression using ultrasmall superparamagnetic iron oxide in Watanabe heritable hyperlipidemic rabbits.

OBJECTIVE: The purpose of this study was to evaluate plaque progression by using MRI with ultrasmall superparamagnetic iron oxide (USPIO) and by histopathological studies. METHODS: We divided 12 Watanabe heritable hyperlipidemic (WHHL) rabbits into 4 groups based on their age (3, 9, 14 and 26 months) and injected them intravenously with 0.8 mmol (Fe) kg(-1) of USPIO (size, 32 nm; concentration, 15 mg dl(-1)). On the fifth post-injection day, they were again given an intravenous injection with 40 µmol kg(-1) of the same USPIO, and MR angiography (MRA) was performed. The signal-to-noise ratio (SNR) in regions of interest in the wall of the upper abdominal aorta was calculated on coronal images. Specimens from the same level of the aorta were subjected to iron staining and RAM-11 immunostaining and used for histopathological study. For statistical analysis of the MRA and histopathological findings, we used analysis of variance [Tukey's honest significant difference (HSD) test]. RESULTS: In 9-month-old rabbits, the SNR was significantly lower than in rabbits of the other ages (p < 0.01), and the area of RAM-11 (DAKO Corporation, Glostrup, Denmark) and iron uptake in the aortic wall was significantly larger (RAM-11, p < 0.01; iron, p < 0.05). These areas were the smallest in 3-month-old rabbits. CONCLUSION: Histopathologically, the number of macrophages was the greatest in 9-month-old rabbits. Our findings indicate that the SNR on MRI scans reflects the number of macrophages in the aortic wall of WHHL rabbits. ADVANCES IN KNOWLEDGE: USPIO-enhanced MRI visualized the accumulation of macrophages in early atherosclerotic plaques of WHHL rabbits in the course of natural progression.

Cerebral atherosclerosis occurs spontaneously in homozygous WHHL rabbits.

We demonstrated that selectively bred homozygous WHHL rabbits known to show hypercholesterolemia and severe coronary atherosclerosis also spontaneously develop cerebral atherosclerosis beginning at 9 months of age. These intracranial lesions occurred in the absence of hypertension in 24 of 25 animals at various sites, mainly along arteries at the base of the brain. No lesions were observed in penetrating arteries. Lesions were rich in smooth muscle cells and fibrous tissue, showing only rare fragmentation or disappearance of the internal elastic lamina, and only limited lipid deposition. Few macrophages were observed in these lesions. No significant correlation was seen between severity of cerebral atherosclerosis and age, systolic blood pressure (BP), serum total cholesterol, or triglyceride concentration. Xanthomas of the pia mater were observed in all 25 rabbits. Arterial findings were similar to those in human cerebral atherosclerosis, indicating that the coronary atherosclerosis-prone homozygous WHHL rabbit represents the first animal model for spontaneous cerebral atherosclerosis.

The effect of selective breeding on the development of coronary atherosclerosis in WHHL rabbits. An animal model for familial hypercholesterolemia.

To increase the incidence of coronary atherosclerosis in Watanabe heritable hyperlipidemic (WHHL) rabbits, an animal model for familial hypercholesterolemia, selective breeding has been carried out. The results obtained are as follows: The incidence of coronary atherosclerosis before selective breeding was 21.8%, whereas that after the breeding was 58.9% (average values for rabbits between 4 and 12 months old), showing that the incidence of coronary atherosclerosis can be increased by an appropriate breeding technique. With respect to the relationships between parents and offspring, if parents had coronary atherosclerosis, then the incidence of the disease in the offspring was 83.3%; therefore the disease was strongly heritable. The mean total cholesterol (TC) and triglyceride (TG) levels of the colony before selective breeding were 579 +/- 163 mg/dl and 471 +/- 144 mg/dl, respectively, while those after the breeding were 665 +/- 151 mg/dl and 603 +/- 178 mg/dl; serum lipid levels increased in parallel with incidence of coronary atherosclerosis. The earliest development of coronary atherosclerosis in the colony after selective breeding was found in 4-month-old rabbits and the whole colony suffered from the disease at 18 months of age.

Inheritability of atherosclerosis and the role of lipoproteins as risk factors in the development of atherosclerosis in WHHL rabbits: risk factors related to coronary atherosclerosis are different from those related to aortic atherosclerosis.

Inheritability of atherosclerosis and the influences of serum lipids on atherosclerosis were examined by following its progression in selectively bred WHHL rabbits. Our studies indicate (1) coronary atherosclerosis is clearly inherited from parents by offspring whereas inheritability of aortic atherosclerosis is uncertain; (2) coronary stenosis is positively correlated to serum cholesterol level, although the correlation coefficient is markedly low: in contrast, no relationship between serum lipid levels and aortic atherosclerosis was observed; (3) cholesterol-rich VLDL showed atherogenicity in aorta, but not in coronary arteries; (4) an unknown lipoprotein detected by 3.6% polyacrylamide gel electrophoresis was related to coronary atherosclerosis, although no relationship between the unknown lipoprotein and aortic atherosclerosis was observed. These findings suggest that there are two types of genetic factors involved in atherosclerosis, one of which is unique to coronary atherosclerosis whereas the other is related to only aortic atherosclerosis.

Fibromuscular cap composition is important for the stability of established atherosclerotic plaques in mature WHHL rabbits treated with statins.

We examined the relationship between plaque vulnerability and fibromuscular cap composition using hydrophilic pravastatin and lipophilic fluvastatin. WHHL rabbits aged 10 months were given pravastatin (50 mg/kg) or fluvastatin (20 mg/kg) for 52 weeks. The atherosclerotic lesions were immunohistochemically or conventionally stained and the components were analyzed with a color image analyzer. Compared with the control group, the plasma cholesterol levels were decreased by about 25% in both statin groups. Pravastatin decreased the lipid components (macrophages+extracellular lipids) in whole aortic plaques by 34% and the fibrous caps of coronary plaques by 55%. Fluvastatin decreased the fibromuscular components (smooth muscle cells+collagen fibers) in whole aortic plaques and in the fibromuscular caps of the aortic and coronary plaques. In the pravastatin group, the vulnerability index, the ratio of (lipid components)/(fibromuscular components), was decreased in whole aortic plaques by 28% and in the fibromuscular caps of coronary lesions by 61%, while the indexes were increased in the fluvastatin group. The incidence of vulnerable plaques was decreased by 74% in the coronary plaques of the pravastatin group. Our results suggest that the stability of atheromatous plaques was improved due to a decrease of the lipid components and vulnerability index of the fibromuscular cap by pravastatin.

Relation of N-glycosylation of apolipoprotein B-100 to cellular metabolism of low density lipoprotein.

We studied the functional role of N-linked sugar chains of apolipoprotein (apo) B-100 of low density lipoprotein (LDL) in cholesterol metabolism. The N-linked sugar chains of apo B-100 of LDL obtained from four homozygous Watanabe heritable hyperlipidemic (WHHL) rabbits were liberated by hydrazinolysis, followed by NaB3H4 reduction and were fractionated by paper electrophoresis and column chromatography. They consisted of one neutral (N) and two acidic (A1, A2) fractions. The ratio of apo B-100 acidic fractions (A1+A2) varied among 4 WHHL rabbits. Serial measurements of serum cholesterol levels showed that they decreased with aging in each of 4 WHHL rabbits. We investigated the relation of the ratio of acidic sugar chains of apo B-100 to the serum cholesterol levels. Reciprocals of the serum cholesterol levels were significantly correlated with the ratio of acidic sugar chains of apo B-100 (r = 0.901, P < 0.001). To elucidate the role of N-linked sugar chains of apo B-100, we investigated cellular uptake of LDL in normal rabbit skin fibroblasts. The amounts of association, degradation and cholesteryl esterification of LDL with a lower ratio of acidic sugar chains at 37 degrees C were greater than those of LDL with a higher ratio of acidic sugar chains. These results suggest that N-glycosylation of apo B-100 may be related with serum cholesterol levels and N-linked sugar chains of apo B-100 may play an important role in cellular metabolism of LDL.

Combination treatment with troglitazone, an insulin action enhancer, and pravastatin, an inhibitor of HMG-CoA reductase, shows a synergistic effect on atherosclerosis of WHHL rabbits.

We examined whether improving insulin resistance augments the antiatherosclerotic effect of LDL reduction. Since WHHL rabbits show hyperinsulinemia and insulin resistance, we administered troglitazone (100 mg/kg), an insulin action enhancer, pravastatin sodium (50 mg/kg), an HMG CoA reductase inhibitor, and a combination of both drugs to 2-month-old WHHL rabbits for 32 weeks. As compared to the control, total cholesterol levels in the plasma and LDL were decreased significantly by 20% in the pravastatin and combination groups. Basal immunoreactive insulin levels and insulin index were decreased significantly by approximately 50% in the troglitazone and combination groups. Surface lesion area of atherosclerosis on the thoracic aorta was decreased significantly by 36% in the combination group and was less in the troglitazone group. Coronary atherosclerosis was decreased significantly by 39% in the combination group and was less in the pravastatin and troglitazone groups. The collagen content in the plaques was decreased in the troglitezone and combination groups and the extracellular lipid deposits were decreased in the pravastatin and combination groups. The incidence and severity of xanthomata in the digital joints were also decreased significantly in the three treated groups. In conclusion, the antiatherogenic effect of the combination treatment is stronger than that of the monotherapy.

Involvement of LDL receptor in the proliferation of vascular smooth muscle cells.

To study the involvement of the low-density lipoprotein (LDL) receptor in the growth of vascular smooth muscle cells (VSMC), we compared the proliferation of cultured VSMC from Watanabe heritable hyperlipidemic (WHHL) rabbits, which lack the LDL receptor, and VSMC from normal Japanese white rabbits in response to platelet derived growth factor (PDGF). The increase in the number of VSMC from WHHL rabbits in response to PDGF (10(-8) M) was significantly lower than that of VSMC from normal rabbits. PDGF stimulated the synthesis of DNA in VSMC from both normal rabbits and WHHL rabbits, but the response was significantly lower in the latter. To determine the involvement of the LDL receptor in the decreased mitogenic response of WHHL rabbit VSMC, we used an anti-LDL receptor monoclonal antibody (MAb) to normal rabbit VSMC; DNA synthesis of VSMC was stimulated by PDGF, but the effect was significantly blocked by the anti-LDL receptor MAb. Mitogen-activated protein (MAP) kinase activity in normal rabbit VSMC was increased by exposure to PDGF, but the effect was significantly suppressed in the presence of the MAb. The anti-LDL receptor MAb markedly inhibited LDL binding to the surface of normal rabbit VSMC. These results suggest that the LDL receptor influences the proliferation of VSMC and thus might be involved in the pathogenesis of atherosclerosis.

Dextran sulfate, a competitive inhibitor for scavenger receptor, prevents the progression of atherosclerosis in Watanabe heritable hyperlipidemic rabbits.

Dextran sulfate competes with binding of modified LDL to the scavenger receptor in macrophages. To elucidate the role of dextran sulfate in the atherosclerotic process, 100 mg of dextran sulfate in drinking water was given to 5 Watanabe heritable hyperlipidemic (WHHL) rabbits for 12 months starting at age 4 months. During the experimental period, there were no significant differences in plasma cholesterol levels between dextran sulfate-treated and untreated rabbits. After 12 months' treatment, accumulation of cholesterol ester in total aorta was significantly suppressed in dextran sulfate-treated rabbits as compared with untreated rabbits (71.4 +/- 22.3 vs. 42.7 +/- 16.5 mg/g dry weight, P < 0.05). Furthermore, lesion area with atherosclerotic plaques in treated rabbits was significantly less than that in untreated rabbits (59.7 +/- 24.5 vs. 30.4 +/- 14.4%, P < 0.05). These results indicate that dextran sulfate might prevent the progression of atherosclerosis by competitively inhibiting the binding of modified LDL to scavenger receptors.