Rivastigmine: an update on therapeutic efficacy in Alzheimer's disease and other conditions.

The results of recent clinical trials with rivastigmine show that in the approved indication of mild to moderate Alzheimer's disease (AD), the drug is effective in the long term. Rivastigmine produces a significant delay in the decline of the three components of AD that have been identified by European guidelines as essential parameters for the assessment of therapeutic efficacy of medicinal products with this indication. These are cognitive function, the ability to perform the usual activities of daily living, and global judgement of the patient's condition by the patient himself, his caregiver and his doctor. Moreover, rivastigmine produces significant control of AD behavioural disorders. This further reduces caregiver burden, reduces the probability of institutionalisation, and enables the reduction or discontinuation of expensive and poorly tolerated antipsychotics. Recent trials also suggest that rivastigmine is effective in moderate to severe Alzheimer's disease, 'mixed' dementia (AD associated with vascular disorders) and Lewy body dementia. Preliminary investigations have also indicated that the drug may provide important benefits in patients with vascular dementia or dementia associated with Parkinson's disease. Pharmacoeconomic studies show that the therapeutic properties of rivastigmine result in economic savings for the care of demented patients living in the community.

Phenotypic expression of familial hypobetalipoproteinemia in three kindreds with mutations of apolipoprotein B gene.

We report the clinical phenotype in three kindreds with familial heterozygous hypobetalipoproteinemia (FHBL) carrying novel truncated apolipoprotein Bs (apoBs) of different sizes (apoB-8.15, apoB-33.4 and apoB-75.7). In D.A. kindred, we found three carriers of a C-deletion in exon 10 leading to the synthesis of apoB-8.15 not detectable in plasma. They showed steatorrhea and fatty liver. In N.L. kindred, the proband is heterozygous for a nonsense mutation in exon 26, leading to the formation of apoB-33.4. He had premature cerebrovascular disease and fatty liver; two apoB-33.4 carriers in this kindred showed only fatty liver. In B.E. kindred, the proband is heterozygous for a T-deletion in exon 26, which converts tyrosine at codon 3435 into a stop codon, resulting in apoB-75.7. The proband, a heavy alcohol drinker, had steatohepatitis, whereas his teetotaller daughter, an apoB-75.7 carrier, had no detectable fatty liver. This study suggests that: i) fatty liver invariably develops in FHBL carriers of short and medium-size truncated apoBs (< apoB-48), but its occurrence needs additional environmental factors in carriers of longer apoB forms; ii) intestinal lipid malabsorption develops only in carriers of short truncated apoBs, which are not secreted into the plasma; and iii) cerebrovascular disease due to premature atherosclerosis may occur even in FHBL subjects.

Plasma protein synthesis in patients with low-grade nephrotic proteinuria.

Overt nephrotic syndrome is characterized by albumin and fibrinogen hyperproduction and reduced very low density lipoprotein apolipoprotein B-100 (VLDL apoB-100) clearance. Whether similar changes also occur in low-grade proteinuria is not known. Thus we measured albumin, fibrinogen, and VLDL apoB-100 kinetics in six patients with modest proteinuria and normal creatinine clearance (P) and in ten control subjects (C) by leucine tracer infusion and precursor-product relationships. In P, plasma albumin concentration was decreased (P < 0.003), whereas concentrations of fibrinogen and VLDL apoB-100 were increased (P < 0.001). In P, albumin fractional secretion rate (FSR) was increased (P < 0.01), fibrinogen FSR was normal, and VLDL apoB-100 FSR was decreased (P < 0.03). As a result, in P, absolute secretion rates (ASR) of albumin and fibrinogen were increased (P < 0.03), whereas VLDL apoB-100 ASR was normal. Albumin FSR was inversely correlated to oncotic pressure in P but not in C. These findings suggest that low-grade nephrotic proteinuria is characterized by simultaneous multiple alterations in turnover rates of albumin, fibrinogen, and VLDL apoB-100. Their pathogenesis, however, appears to be multifactorial.

Changes in HCV viremia following LDL apheresis in a HCV positive patient with familial hypercholesterolemia.

It has been suggested that hepatitis C virus (HCV) can be associated with beta-lipoprotein in human serum. According to this, the LDL receptor could promote endocytosis of such a virus. In the present study, we evaluated the changes in HCV viremia in a HCV positive patient with familial hypercholesterolemia, undergoing both selective (DALI System, Fresenius) and non-selective (plasma exchange) LDL apheresis. HCV-RNA levels did not decrease following selective LDL apheresis, on the contrary showed a random, odd variation pattern (from -35% to +72%). Conversely, plasma exchange steadily induced a drop in HCV viremia (-35/43%), to a lower extent than that of a totally intravascular plasmaprotein, i.e., alpha 2-macroglobulin (-53/54%). These data indicate that beta-lipoprotein may not function as a plasma carrier of HCV, at least in the present case. Moreover, a continuous, quantitatively unforeseeable circulation of HCV virions from the intravascular plasma compartment to other extravascular and intracellular sites, seems to occur during an apheresis session.

Lipoprotein(a) and lipoprotein profile in healthy centenarians: a reappraisal of vascular risk factors.

In this study we assessed whether widely accepted risk factors for atherosclerotic vascular diseases such as lipoprotein(a) [Lp(a)], a cholesterol-rich lipoprotein under strict genetical control, and other lipid parameters change with age. The variations of blood levels and the pathophysiological role of Lp(a) in old people, and particularly in the oldest old, are unknown. Accordingly, we measured Lp(a) levels as well as total, LDL, and HDL cholesterol (CT), and triglycerides (TG) in sera from 75 healthy centenarians, 114 randomly selected subjects under 65 years, 73 randomly selected elderly people, and 30 healthy selected elderly people. The results showed that Lp(a) serum levels did not vary by age group, including centenarians. Remarkably, one-quarter of the centenarians had high Lp(a) serum levels even though they never suffered from atherosclerosis-related diseases. At variance with young and aged people, centenarians with high Lp(a) serum levels also had high plasma concentrations of the proinflammatory cytokine IL-6, suggesting that genetic control of the Lp(a) serum level may attenuate with age and that environmental factors such as chronic subclinical inflammatory processes may play a role. We also showed that most centenarians are paradoxically characterized by low HDL-CT and relatively high TG levels, which together are considered to be strong risk factors for coronary heart disease. On the whole, these data support the hypothesis that a continuous and complex reshaping of lipid metabolism occurs in physiological aging, likely contributing to successful aging.

Homozygous familial hypobetalipoproteinemia. Increased LDL catabolism in hypobetalipoproteinemia due to a truncated apolipoprotein B species, apo B-87Padova.

Mutations on the apolipoprotein (apo) B gene that interfere with the full-length translation of the apoB molecule are associated with familial hypobetalipoproteinemia (FHBL), a disease characterized by the reduction of plasma apoB and LDL cholesterol. In this report, we describe an FHBL kindred carrying a unique truncated apoB form, apoB-87Padova. Sequence analysis of amplified genomic DNA identified a single G deletion at nucleotide 12032, which shifts the translation reading frame and causes a termination at amino acid 3978. Two homozygous subjects and seven heterozygous relatives were studied. Although homozygous individuals had only trace amounts of LDL, they were virtually free from the symptoms typical of homozygous FHBL subjects. We investigated the in vivo turnover of radiolabeled normal apoB-100 LDL and apoB-87 LDL in one homozygous patient and two normal control subjects. ApoB-87 LDL showed a similar metabolism in all three subjects, with a fractional catabolic rate more than double that of normal LDL. The rate of entry of apoB-87 in the LDL compartment was also markedly decreased compared with normal apoB-100. The increased in vivo catabolism of apoB-87 LDL was paralleled in vitro by a 2.5-fold increased ability of these particles to inhibit the uptake and degradation of normal apoB-100 LDL by normal human cultured fibroblasts. These results indicate that apoB-87 LDL has an enhanced ability to interact with the LDL receptor, the increased apoB catabolism contributes to the hypobetalipoproteinemia and may explain the mild expression of the disease in the two homozygous individuals.

Heterozygous apolipoprotein C-II deficiency: lipoprotein and apoprotein phenotype and RsaI restriction enzyme polymorphism in the Apo C-IIPadova kindred.

Deficiency of apolipoprotein C-II (apo C-II), the cofactor for lipoprotein lipase, results in the familial chylomicronaemia syndrome characterized by severe hypertriglyceridaemia and fasting chylomicronaemia. To investigate the biochemical features of the heterozygous state for apo C-II deficiency, we characterized the lipid, lipoprotein and apolipoprotein profiles in 18 relatives of two affected individuals (brother and sister) homozygous for the apo C-IIPadova gene defect which results in the synthesis of a truncated 36 amino acid apolipoprotein. Carrier status was established in first degree relatives as well as in seven non-obligate heterozygotes by restriction enzyme analysis of amplified apo C-II genomic DNA using RsaI. No significant differences in lipid, lipoprotein and apo C-II levels were observed in heterozygotes when compared to unaffected family members. Thus, in this study, the carrier state was not associated with hypertriglyceridaemia or reduced plasma levels of apo C-II. However, analysis of amplified DNA from members of the apo C-IIPadova kindred by digestion with the enzyme RsaI, which identifies the mutant apo C-II, permitted the identification of heterozygous family members which could not be recognized by measuring either fasting triglycerides or plasma apo C-II levels. This study provides further evidence that apo C-II deficiency syndrome is a heterogeneous disease not only at the molecular level but also on the clinical ground with variable phenotypic expression in heterozygous individuals from different kindreds.

Acute effects of HMG-CoA reductase inhibitors on biliary lipids in patients with interrupted enterohepatic circulation.

HMG-CoA reductase inhibitors decrease serum cholesterol by inhibiting hepatic cholesterol synthesis, but their influence on biliary lipids is not well characterized. In the present study Pravastatin (80 mg) was administered as a single oral dose to 10 patients with external bile fistula, after 1 week of interruption of the enterohepatic circulation, in order to assess the effect of inhibition of hepatic cholesterol synthesis on biliary lipids in conditions of stimulated bile acid synthesis. Bile was collected every hour for 12 h. On the day before, the same procedure was applied with a placebo, and collected bile used as control. Pravastatin decreased both bile acid and phospholipid concentration to about 60% of basal values; this change was still significant after 10 h. Cholesterol concentration was also decreased to about 70% of basal values, but this change was significant only from the 5th to the 7th h. The per cent of cholic and chenodeoxycholic acid was not affected by the drug, but the ratio of glyco- to tauroconjugated bile acids was decreased to about half the initial values. Bilirubin concentration exhibited a late increase, suggesting a reduction in the bile flow. These results suggest that, in patients with interrupted enterohepatic circulation, biliary excretion of bile acids can be largely dependent on hepatic cholesterol synthesis.

A nonsense mutation in the apolipoprotein C-IIPadova gene in a patient with apolipoprotein C-II deficiency.

The apo C-II gene from a patient with apo C-II deficiency has been sequenced after amplification by the polymerase chain reaction. A substitution of an adenosine for a guanosine at position 3002 in exon 3 of the patient's gene was identified by sequence analysis. This mutation leads to the introduction of a premature termination codon (TAA) at a position corresponding to amino acid 37 of mature apo C-II and to the formation of a new Rsa I restriction enzyme site not present in the normal apo C-II gene. Amplification of DNA from family members by the polymerase chain reaction and digestion with Rsa I established that the patient is a true homozygote for the mutation. Analysis of the patient's plasma by two-dimensional gel electrophoresis and immunoblotting detected an apo C-II that exhibited abnormal electrophoretic mobility. We propose that the C to A substitution in the apo C-IIPadova gene is the primary genetic defect that leads to premature termination and the synthesis of a truncated 36 amino acid apo C-II that is unable to activate lipoprotein lipase.

Apolipoprotein C-II deficiency: identification of a structural variant ApoC-II Padova.

Apolipoprotein(apo) C-II DNA, RNA and protein from a patient with a familial deficiency of apoC-II were evaluated and compared to normal individuals. No major defect of the apoC-II gene could be detected by Southern blot hybridization. Northern and slot blot analyses of total liver RNA documented normal levels of a normal sized apoC-II mRNA. Immunohistochemical studies of the liver of the apoC-II deficient patient revealed a normal to slightly elevated intracellular content of the C-II apolipoprotein. Plasma apoC-II was 3 to 5% of normal apoC-II levels and exhibited abnormal electrophoretic mobility on two dimensional gel electrophoresis and immunoblotting. We postulate that at the molecular level, the deficiency of apoC-II in the plasma of this patient results from a structural defect in the coding portion of the apoC-II gene leading to either defective secretion of cellular apoC-II or increased catabolism of a structurally defective apoC-II in plasma.

Postnatal transformations of alveolar surfactant in the rabbit: changes in pool size, pool morphology and isoforms of the 32-38 kDa apolipoprotein.

To clarify perinatal transformations of surfactant we performed lung lavage in term fetuses and in 0-24-h-old newborn rabbits. Lavage fluid was separated into three pools, namely lavage pellet, lavage supernatant and cells. We found that at birth the pellet contains 94.1 +/- 1.4% (S.E.) saturated phosphatidylcholine, while the supernatant and cells contain traces of it. At birth the pellet contains secreted lamellar bodies while the supernatant lacks any recognizable structure. After birth, the alveolar saturated phosphatidylcholine level increases 5.1-times in 24 h, the proportions between pools reaching adult values in 90 min (pellet = 75.9 + 4.8%, supernatant = 22.7 +/- 4.9%), and small vesicles appear in the supernatant, probably originating from the turnover of alveolar surfactant during breathing. The saturated phosphatidylcholine associated with cells remains unchanged. At birth, the 32-38 kDa surfactant apolipoprotein appears to be less extensively sialylated than in adult life.

Long term-effect of fenofibrate on lipoprotein level and composition in different types of genetic hyperlipidemias.

Fenofibrate effect on plasma lipids and lipoproteins was studied in 23 patients with primary hyperlipoproteinemias (HLP): 12 with familial hypercholesterolemia (FH), 5 with combined HLP and 6 with Type III HLP. Trial lasted from 8 to 10 months. Cholesterol and triglycerides were significantly reduced in all patients as follows: cholesterol dropped 26% in FH (p less than 0.001), 32% in combined HLP and 48% in type III HLP; triglycerides dropped 29%, 64% and 72% respectively. This drop involved LDL in FH, VLDL and LDL in combined HLP and beta-VLDL in type III HLP patients. In FH and combined HLP patients we observed a +10% (p less than 0.01) and a +9% (n.s.) increase of HDL, respectively. Two patients had a mild SGOT and SGPT increase and three had a vescicular cutaneous erythema.

Lipoprotein modifications during dietary treatment in patients with primary type V hyperlipoproteinaemia.

Type V hyperlipoproteinaemia is a disorder of lipid transport characterized by the accumulation in serum of chylomicrons and very low density lipoproteins. The purpose of the study was the analysis of serum lipids and lipoproteins by ultracentrifugation in nine patients with primary type V hyperlipoproteinaemia before and during dietary treatment. After 30 days of balanced isocaloric diet mean serum triglycerides fell from 25.4 +/- 15.0 (mean +/- SD) to 2.8 +/- 1.7 mmol l-1. At the same time the chylomicrons and the very low density lipoproteins of flotation rate higher than 100 disappeared from the serum while the remaining very low density lipoproteins maintained unaltered their normal protein-lipid composition. After 30 days the low density lipoproteins increased significantly in concentration (from 1.6 +/- 0.8 to 4.1 +/- 1.1 mmol l-1 cholesterol) and their percentage content of cholesterol and triglyceride was increased and reduced, respectively. The highest concentration of intermediate density lipoprotein cholesterol was observed after 15 days of treatment (1.2 +/- 0.6 mmol l-1. The abnormally low concentrations and the physicochemical properties of the high density lipoproteins remained unchanged throughout the study (from 0.6 +/- 0.2 to 0.8 +/- 0.2 mmol l-1 cholesterol concentration) and no high density lipoproteins two (HDL2) were observed at any time. The effects of this treatment were an increase in low density and marginal change in high density lipoproteins which are considered, respectively, a positive and a negative risk factor for atherosclerosis.

Separation of apolipoprotein B species by agarose-acrylamide gel electrophoresis.

Human apolipoprotein (apo) B has been recognized to exist in two different forms designated apoB-100 and apoB-48. The two apoB forms are usually separated by NaDodSO4 gel electrophoresis with a low percentage polyacrylamide gel in a tube gel apparatus. However, the matrix of this low percentage gel is relatively weak, and one can separate the two forms of apoB in a slab gel apparatus only if one utilizes a gradient polyacrylamide gel or a higher percentage polyacrylamide gel which results in a poorer separation of the protein bands. We have developed an agarose-acrylamide gel electrophoretic method to separate the two major apoB forms. The gel is a mixture of 0.5% agarose and 2% acrylamide. The agarose-acrylamide method is fast, has the advantage of being able to be used on an analytical or preparative scale in a vertical slab gel apparatus, and the gel is of sufficient strength to be used in immunoblotting and/or radioautography.

Abnormal low density lipoprotein metabolism in apolipoprotein E deficiency.

Apolipoprotein(apo) E deficiency is an inherited disease characterized by type III hyperlipoproteinemia and less than 1% normal plasma apoE concentration. The role of apoE in LDL metabolism was investigated by quantitating the metabolism of radiolabeled normal and apoE-deficient LDL in both normal and apoE-deficient subjects. ApoE deficiency resulted in an accumulation of plasma IDL, and a decreased synthesis of LDL consistent with a block in the conversion of IDL to LDL. The LDL isolated from the apoE-deficient patient was similar to normal LDL in hydrated density, size, and composition. However, the apoE-deficient LDL was kinetically abnormal with delayed catabolism in both normal subjects and the apoE-deficient patient. In addition, the catabolism of normal LDL in the apoE-deficient subject was increased. These results were interpreted as indicating that apoE is necessary for the conversion of IDL to LDL and the formation of kinetically normal LDL. The rapid catabolism of normal LDL in the apoE-deficient patient suggests an up-regulation of the hepatic LDL receptor pathway. Based on these results, apoE is proposed to play an important role in the conversion of IDL to LDL, the formation of kinetically normal LDL, and the regulation of LDL receptor function.

Apolipoprotein C-II deficiency syndrome. Clinical features, lipoprotein characterization, lipase activity, and correction of hypertriglyceridemia after apolipoprotein C-II administration in two affected patients.

Two patients (brother and sister, 41 and 39 yr of age, respectively) have been shown to have marked elevation of plasma triglycerides and chylomicrons, decreased low density lipoproteins (LDL) and high density lipoproteins (HDL), a type I lipoprotein phenotype, and a deficiency of plasma apolipoprotein C-II (apo C-II). The male patient had a history of recurrent bouts of abdominal pain often accompanied by eruptive xanthomas. The female subject, identified by family screening, was asymptomatic. Hepatosplenomegaly was present in both subjects. Analytical and zonal ultracentrifugation revealed a marked increase in triglyceride-rich lipoproteins including chylomicrons and very low density lipoproteins, a reduction in LDL, and the presence of virtually only the HDL3 subfraction. LDL were heterogeneous with the major subfraction of a higher hydrated density than that observed in plasma lipoproteins of normal subjects. Apo C-II levels, quantitated by radioimmunoassay, were 0.13 mg/dl and 0.12 mg/dl, in the male and female proband, respectively. A variant of apo C-II (apo C-IIPadova) with lower apparent molecular weight and more acidic isoelectric point was identified in both probands by two-dimensional gel electrophoresis. The marked hypertriglyceridemia and elevation of triglyceride-rich lipoproteins were corrected by the infusion of normal plasma or the injection of a biologically active synthesized 44-79 amino acid residue peptide fragment of apo C-II. The reduction in plasma triglycerides after the injection of the synthetic apo C-II peptide persisted for 13-20 d. These results definitively established that the dyslipoproteinemia in this syndrome is due to a deficiency of normal apo C-II. A possible therapeutic role for replacement therapy of apo C-II by synthetic or recombinant apo C-II in those patients with severe hypertriglyceridemia and recurrent pancreatitis may be possible in the future.

Improvement of electroimmunoassay for apolipoproteins B and A-I. (Preliminary observations in postprandial phase).

A reliable method for determining serum apoprotein levels is an essential condition for investigating the role of apoproteins in atherogenesis. Electroimmunoassay according to Laurell has been studied and applied with some modifications for the determination of the two main apoproteins: Apo A-I and Apo B. Apo B immunoplates containing 0.4% v/v of rabbit anti-Apo B antiserum were processed for 4 h at 10 V/cm. Samples were incubated at 52 degrees C for 3 h, diluted and then 10 microliter were seeded in each well. Apo A-I immunoplates (8% v/v of sheep antiserum) were processed for 24 h at 2 V/cm. Agarose gel concentration, exsiccation procedure and staining were the same for both apoproteins. The method was standardized employing a secondary standard consisting of a serum pool obtained from normal subjects. Apo A-I and Apo B levels of the pool have been previously determined employing as primary standards the HDL3 (1.120-1.230 g/ml) and the LP-B (1.035-1.050 g/ml) fractions, respectively, which were isolated by preparative ultracentrifugation. Preliminary observations from a study on 20 healthy volunteers with normal lipid levels revealed different apoprotein levels in young men and women and significant differences between postmenopausal women and women in the fertile age.