Biomedical subjects

M Averna

Publications and source records attributed to M Averna.

At least 19 recordsLinked to original sources

PubMed2003

Changes in calpastatin localization and expression during calpain activation: a new mechanism for the regulation of intracellular Ca(2+)-dependent proteolysis.

The amount of calpastatin directly available in cytosol is under the control of [Ca2+] and [cyclic AMP]. Prolonged calpain activation also promotes degradation of calpastatin. The fluctuation of calpastatin concentration in cell soluble fraction is accompanied by an initial decrease in calpastatin gene expression, followed by a fivefold increase in its expression when the inhibitor protein is degraded. This process can be conceptualized as a mechanism to regulate calpastatin availability in the cell. This conclusion is supported by the fact that calpain, the other component of this proteolytic system, undergoes changes in its levels of expression in a much more limited manner. Furthermore, this process can be observed both in cells exposed to different natural stimuli, or in other cell lines. Modification of calpastatin gene expression might represent a new tool for the in vivo control of the regulatory machinery required for the modulation of Ca(2+)-dependent proteolysis.

Calcium↗

PubMed2003

Ventricular arrhythmias in hypertensive patients with mild metabolic alterations.

BACKGROUND: Ventricular arrhythmias (VA) are prevalent in hypertensives, as well as in diabetics and dyslipidemics. The aim of our study is to evaluate if alterations in glucose or lipid metabolism increase the risk of VA in hypertensive patients. METHODS: Sixty-eight patients (24 males, 44 females, aged 45-78), with mild-moderate hypertension, were divided into 2 groups: group A (n=24) normoglycemic and normolipemic, and group B (n=44) with serum cholesterol >220 mg% and/or triglycerides >165 mg% and/or glucose >110 mg%. Patients with clinical or ECG signs of ischemia or kaliemia <3.5 mEq/l, were not admitted to the study. After a 15-days pharmacological wash-out, 24-hour Holter ECG and echocardiography were performed on all patients. RESULTS: Ventricular ectopic beats (VEB) were found in 66% of group A and 100% of group B (difference =34%, 95% C.I. =18-50%). VEB were less severe in group A (Lown's class 1-2), while group B had VEB ranging from class 1 to 5. Left ventricular hypertrophy (LVH) was found in 33% of group A and in 72.7% of group B, and was not closely correlated to VA. Group B patients with LVH had more severe VA (4 A, 4 B and 5 Lown's class). CONCLUSIONS: Hyperglycemia and/or dyslipidemia, even mild, seem to increase the risk of VA in the hypertensive patient with or without LVH. It suggests that 24 hour Holter ECG may be useful to select, among patients with metabolic alterations, those at high risk for VA.

Aged↗

PubMed2001

Age-dependent degradation of calpastatin in kidney of hypertensive rats.

Hypertensive rats from the Milan strain show a significant decrease in calpastatin activity as compared with normotensive control animals. Calpastatin deficiency is age-related and highly relevant in kidney, heart, and erythrocytes and of minor entity in brain tissue. In normotensives the changes during aging in the levels of calpastatin activity and mRNA are consistent with an increase of calpastatin protein. In hypertensive rats such a relationship during aging is not observed, because a progressive accumulation of mRNA is accompanied by a lower amount of calpastatin protein as compared with control rats. Together with the low level of calpastatin in kidney of hypertensive rats, a progressive accumulation of an active 15-kDa calpastatin fragment, previously shown to represent a typical product of calpain-mediated calpastatin degradation, is also observed. Evidence for such intracellular proteolysis by Ca(2+)-activated calpain is provided by the normalization of the calpastatin level, up to that of control animals, in hypertensive rats treated with drugs known to reduce both blood pressure and intracellular Ca(2+) influx. Further evidence is provided by the disappearance, in these conditions, of the 15-kDa calpastatin fragment. These data allow the conclusion that calpastatin degradation is a relevant part of the overall mechanism for regulating calpain activity.

Age Factors↗

PubMed2001

Changes in intracellular calpastatin localization are mediated by reversible phosphorylation.

We have previously reported that, in neuroblastoma LAN-5 cells, calpastatin is in an aggregated state, close to the cell nucleus [de Tullio, Passalacqua, Averna, Salamino, Melloni and Pontremoli (1999) Biochem. J. 343, 467-472]. In the present paper, we demonstrate that aggregated calpastatin is predominantly in a phosphorylated state. An increase in intracellular free [Ca2+] induces both dephosphorylation of calpastatin, through the action of a phosphoprotein phosphatase, and its redistribution as a soluble inhibitor species. cAMP, but not PMA-induced phosphorylation, reverses calpastatin distribution favouring its aggregation. This intracellular reversible mechanism, regulating the level of cytosolic calpastatin, could be considered a strategy through which calpain can escape calpastatin inhibition, especially during earlier steps of its activation process.

Calcium-Binding Proteins↗

PubMed2001

Dietary cholate increases plasma levels of apolipoprotein B in mice by posttranscriptional mechanisms.

To induce atherogenesis in mice, a high fat (HF) diet is supplemented with cholic acid (CA), which increases apoB-containing particles and lower apoA-I-containing particles. HF diet without CA increases levels of both HDL and LDL, suggesting that CA may be responsible for the elevation of LDL and lowering of HDL. The mechanism of dietary CA-induced lowering of apoA-I-containing particles has recently been reported. In this study, we examined the mechanism of CA- and HF-induced elevation of apoB-containing lipoproteins in mice. Mice were fed the following four diets: control chow (C), high fat high cholesterol, (HF), control and 0.5% cholate (CA), and HF+CA. Dietary CA increased the plasma levels of apoB-containing particles by approximately 2-fold when compared to control; VLDL levels increased 2-fold, and LDL levels increased 1.3-fold. On HF diet, VLDL increased by 1.4-fold, and LDL by 2-fold, suggesting that CA and HF-induced increases of apoB-containing particles occurred by different mechanisms. We investigated the potential mechanisms regulating plasma levels of apoB in CA- and HF-fed mice. Although hepatic apoB mRNA levels did not change on CA diet, apoB-100 mRNA increased relative to B-48 as a result of decreased editing of apoB mRNA. Measurements of hepatic LDL receptor mRNA suggested that CA diet down-regulated LDL receptor mRNA, possibly by increasing the levels of hepatic cholesterol. Since plasma and hepatic vitamin E levels did not show significant changes on CA-containing diets, it suggests that dietary CA did not act by increasing the absorption of dietary fat. Hepatic lipase, known to modulate plasma levels of apoB-containing particles, did not show changes in CA- or HF-fed mice. Taken together, these results suggest that dietary CA increased apoB-containing particles both in chow-fed and fat-fed mice by enhancing the relative production of apoB-100, and also by reducing LDL receptor-mediated clearance of apoB-containing particles. Thus, dietary cholate modulates plasma levels of apoB primarily by posttranscriptional mechanisms.

Animals↗

PubMed2001

Estrogen increases hepatic lipase levels in inbred strains of mice: a possible mechanism for estrogen-dependent lowering of high density lipoprotein.

We have shown mouse to be an useful animal model for studies on the estrogen-mediated synthesis and secretion of lipoproteins since, unlike in rats, low density lipoprotein receptors are not upregulated in mice. This results into the elevation of plasma levels of apolipoprotein (apo) B and apoE, and lowering of apoA-1-containing particles. The mechanisms of apoB and apoE elevation by estrogen have been elucidated, but the mechanism of lowering of plasma levels of HDL is still not known. Among other factors, apoA-I, cholesterol ester transfer protein (CETP), scavenger receptor B1 (SR-B1), and hepatic lipase are potential candidates that modulate plasma levels of HDL. Since estrogen treatment increased hepatic apoA-I mRNA and apoA-I synthesis, and mouse express undetectable levels of CETP, we tested the hypothesis that estradiol-mediated lowering of HDL in mice may occur through modulation of hepatic lipase (HL). Four mouse strains (C57L, C57BL, BALB, C3H) were administered supraphysiological doses of estradiol, and plasma levels of HDL as well as HL mRNA were quantitated. In all 4 strains estradiol decreased plasma levels of HDL by 30%, and increased HL mRNA 2-3 fold. In a separate experiment groups of male C57BL mouse were castrated or sham-operated, and low and high doses of estradiol administered. We found 1.4-2.5 fold elevation of HL mRNA with concomitant lowering of HDL levels. Ten other mouse strains examined also showed estradiol-induced elevation of HL mRNA, but the extent of elevation was found to be strain-specific. Based on these studies, we conclude that hepatic lipase is an important determinant of plasma levels of HDL and that HL mRNA is modulated by estrogen which in turn may participate in the lowering of plasma levels of HDL.

Animals↗

PubMed2001

Determinants of enhanced thromboxane biosynthesis in renal transplantation.

BACKGROUND: Despite great improvement in patient and graft survival, the long-term morbidity and mortality in renal transplant recipients (RTRs) are still significant, with a high incidence of cardiovascular disease-related deaths. METHODS: We investigated thromboxane (TXA2) biosynthesis and endothelial and coagulative activation in 65 patients who received a renal transplant. RESULTS: The rate of TXA2 biosynthesis (urinary 11-dehydro-TXB2 excretion largely reflects platelet TXA2 production in vivo) was significantly (P < 0.0001) higher in RTRs than in healthy subjects. Plasma von Willebrand factor (vWF) and thrombin-antithrombin (TAT) complexes were significantly higher (P < 0.001) in RTRs compared with controls. Urinary 11-dehydro-TXB2 directly correlated with plasma vWF and cholesterol. We next examined the relative influence of cyclosporine A (CsA) on TXA2 biosynthesis and endothelial activation, comparing a group of RTRs not receiving CsA with an age- and sex-matched group of patients treated with CsA. Urinary excretion of 11-dehydro-TXB2 and plasma levels of vWF were significantly increased in RTRs who received CsA compared with those who did not. After an overall follow-up of 120 months, RTRs who experienced cardiovascular events had a higher frequency of abnormal plasma levels of vWF than patients who remained event free. CONCLUSION: Renal transplantation is associated with in vivo platelet activation highly related to endothelial activation. This is particularly evident in CsA-treated patients. Administration of drugs that are able to reduce or eliminate thromboxane-dependent platelet activation in vivo may be beneficial to reduce the risk of cardiovascular events in RTRs.

Adult↗

PubMed2000

Differential degradation of calpastatin by mu- and m-calpain in Ca(2+)-enriched human neuroblastoma LAN-5 cells.

In neuroblastoma LAN-5 cells during calpain activation, in addition to the two expressed 70 kDa and 30 kDa calpastatin forms, other inhibitory species are produced, having molecular masses of 50 kDa and 15 kDa. At longer times of incubation, both native and new calpastatin species disappear. The formation of these new calpastatins as well as the decrease in intracellular total calpastatin activity are mediated by calpain itself, as indicated by the effect of the synthetic calpain inhibitor I, which prevents both degradative processes. Analysis of the calcium concentrations required for the two processes indicates that the first conservative proteolytic event is mediated by micro-calpain, whereas the second one is preferentially carried out by m-calpain. The appearance of the 15 kDa form, containing only the calpastatin repetitive inhibitory domain and identified also in red cells of hypertensive rats as the major inhibitor form, can be considered a marker of intracellular calpain activation, and it can be used for the monitoring of the involvement of calpain in pathological situations.

Animals↗

PubMed2000

Acyl-CoA-binding protein is a potent m-calpain activator.

Acyl-CoA-binding protein, a 20-kDa homodimer that exerts many physiological functions, promotes activation of the classic calpain forms, most markedly that of the m-isozyme. This protein factor was purified from rat skeletal muscle and was also expressed in Escherichia coli. Both native and recombinant acyl-CoA-binding proteins show the same molecular properties and an identical capacity to decrease the [Ca(2+)] required for m-calpain activity. The binding of long-chain acyl-CoAs to acyl-CoA-binding protein does not modify the activating effect on calpains. Acyl-CoA-binding protein seems to be involved in the m-calpain regulation process, whereas the previously identified UK114 activator is a specific modulator of micro-calpain. Acyl-CoA-binding protein is proposed as a new component of the Ca(2+)-dependent proteolytic system. A comparative analysis among levels of classic calpains and their activator proteins is also reported.

Amino Acid Sequence↗

PubMed2000

Dietary cholic acid lowers plasma levels of mouse and human apolipoprotein A-I primarily via a transcriptional mechanism.

To induce dietary atherosclerosis in mice, high-fat/high-cholesterol (HF) diets are frequently supplemented with cholic acid (CA). This diet produces low plasma levels of high-density lipoprotein (HDL) and high levels of low-density lipoprotein (LDL). However, HF diets without any added CA, which more closely resemble human diets, increase levels of both HDL and LDL, suggesting that CA may be responsible for the lowering of HDL. Our aim was to examine the potential mechanism responsible for the lowering of HDL. Nontransgenic (NTg) C57BL mice and apoA-I-transgenic (apoAI-Tg) mice, with greatly increased basal apoA-I and HDL levels, were used. Mice were fed the following four diets: control (C), high-fat/high-cholesterol (HF), control and 1% cholate (CA) and HF + CA. Dietary CA reduced plasma HDL levels by 35% in NTg and 250% in apoAI-Tg mice, independent of the fat or cholesterol content of the diet. Hepatic apoA-I mRNA decreased 30% in NTg and 180% in apoAI-Tg mice. Hepatic apoA-I synthesis and apoA-I mRNA transcription rates also decreased in parallel with apoA-I mRNA levels, suggesting that the CA-induced decreases in plasma apoA-I levels occurred primarily via decreasing apoA-I mRNA transcription rates. An HF diet increased HDL levels 1.8-fold in NTg and 1.5-fold in apoAI-Tg mice. Addition of CA to the HF diet lowered HDL levels by 1.6-fold in NTg and 2. 5-fold in apoAI-Tg mice. Transfection studies with the apoA-I promoter suggested the presence of a putative cis-acting element responsible for the CA-mediated down-regulation of the apoA-I promoter activity. Measurements of apoA-I regulatory protein-1 (ARP-1) mRNA, a negative regulator of the apoA-I gene in the mouse liver showed that CA increased the ARP-1 mRNA levels. Because apoA-I gene transcription alone was not sufficient to account for the lowering of plasma HDL levels, scavenger receptor-B1 (SR-B1) and hepatic lipase (HL) mRNAs levels were quantitated. The levels of SR-B1 and HL mRNA were not changed by dietary CA. These studies suggest that dietary cholate regulates plasma levels of apoA-I primarily by a transcriptional mechanism via a putative bile acid response element involving a negative regulator of apoA-I, and partly by an unidentified post-transcriptional mechanism.

Animals↗

PubMed2000

Clinical expression of familial hypercholesterolemia in clusters of mutations of the LDL receptor gene that cause a receptor-defective or receptor-negative phenotype.

Seventy-one mutations of the low density lipoprotein (LDL) receptor gene were identified in 282 unrelated Italian familial hypercholesterolemia (FH) heterozygotes. By extending genotype analysis to families of the index cases, we identified 12 mutation clusters and localized them in specific areas of Italy. To evaluate the impact of these mutations on the clinical expression of FH, the clusters were separated into 2 groups: receptor-defective and receptor-negative, according to the LDL receptor defect caused by each mutation. These 2 groups were comparable in terms of the patients' age, sex distribution, body mass index, arterial hypertension, and smoking status. In receptor-negative subjects, LDL cholesterol was higher (+18%) and high density lipoprotein cholesterol lower (-5%) than the values found in receptor-defective subjects. The prevalence of tendon xanthomas and coronary artery disease (CAD) was 2-fold higher in receptor-negative subjects. In patients >30 years of age in both groups, the presence of CAD was related to age, arterial hypertension, previous smoking, and LDL cholesterol level. Independent contributors to CAD in the receptor-defective subjects were male sex, arterial hypertension, and LDL cholesterol level; in the receptor-negative subjects, the first 2 variables were strong predictors of CAD, whereas the LDL cholesterol level had a lower impact than in receptor-defective subjects. Overall, in receptor-negative subjects, the risk of CAD was 2.6-fold that of receptor-defective subjects. Wide interindividual variability in LDL cholesterol levels was found in each cluster. Apolipoprotein E genotype analysis showed a lowering effect of the epsilon2 allele and a raising effect of the epsilon4 allele on the LDL cholesterol level in both groups; however, the apolipoprotein E genotype accounted for only 4% of the variation in LDL cholesterol. Haplotype analysis showed that all families of the major clusters shared the same intragenic haplotype cosegregating with the mutation, thus suggesting the presence of common ancestors.

Adult↗

PubMed1999

Changes in intracellular localization of calpastatin during calpain activation.

Localization of the two main components of the Ca(2+)-dependent proteolytic system has been investigated in human neuroblastoma LAN-5 cells. Using a monoclonal antibody which recognizes the N-terminal calpastatin domain, it has been shown that this inhibitory protein is almost completely confined in two granule-like structures not surrounded by membranes. Similar calpastatin distribution has been found in other human and in murine cell types, indicating that aggregation of calpastatin is a general property and not an exclusive characteristic of neuronal-like cells. The existence of such calpastatin aggregates is confirmed by the kinetics of calpastatin-activity release during rat liver homogenization, which does not correspond to the rate of appearance of cytosolic proteins or to the disruption of membrane-surrounded organelles. Calpastatin distribution is affected by the intracellular increase in free Ca(2+), which results in calpastatin progressively becoming a soluble protein. However, calpain is distributed in the soluble cell fraction and, in activating conditions, partially accumulates on the plasma membrane. Similar behaviour has been observed in calpastatin localization in LAN-5 cells induced with retinoic acid, suggesting that the proteolytic system is activated during the differentiation process of these cells. The involvement of calpastatin in controlling calpain activity, rather than its activation process, and the utilization of changes in calpastatin localization as a marker of activation of the system is discussed.

Animals↗

PubMed1999

Phosphorylation of rat brain calpastatins by protein kinase C.

Calpastatin, the natural inhibitor of calpain, is present in rat brain in multiple forms, having different molecular masses, due to the presence of one (low Mr form) or four (high Mr form) repetitive inhibitory domains. Recombinant and native calpastatin forms are substrates of protein kinase C, which phosphorylates a single serine residue at their N-terminus. Furthermore, both low and high Mr calpastatins are phosphorylated by protein kinase C at the same site. These calpastatin forms are phosphorylated also by protein kinase A, although with a lower efficiency. The incorporation of a phosphate group determines an increase in the concentration of Ca2+ required to induce the formation of the calpain-calpastatin complex. This effect results in a large decrease of the inhibitory efficiency of calpastatins. We suggest that phosphorylation of calpastatin represents a mechanism capable to balance the actual amount of active calpastatin to the level of calpain to be activated.

Animals↗

PubMed1999

Rapid screening of the LDL receptor point mutation FH-Genoa/Palermo. Mutation in brief no. 238. Online.

The LDL-receptor gene point mutation FH-Genoa/Palermo is the most frequent mutation responsible for Familial Hypercholesterolemia in Sicily. The mutation does not introduce or abolish any useful restriction site. We establish a GeneComb-based strategy to identify this mutation in a population of Sicilian unrelated clinically diagnosed FH probands. The method was very sensitive and specific; 12 out of 90 (13.3%) unrelated FH probands were found to carry the FH-Genoa/Palermo mutation. According to these results, the FH-Genoa/Palermo is the more frequent LDL-receptor gene mutation among the Sicilian FH patients. Moreover FH-Genoa/Palermo is the mutation cluster to date more represented in Southern Italy.

Genetic Testing↗

PubMed1999

Regulation of the apolipoprotein B in heterozygous hypobetalipoproteinemic knock-out mice expressing truncated apoB, B81. Low production and enhanced clearance of apoB cause low levels of apoB.

Low levels of cholesterol are protective against development of coronary artery disease. Heterozygous hypobetalipoproteinemic individuals expressing truncated apolipoprotein (apo)B as a result of mutation in the apob gene have low levels of cholesterol and apoB in their plasma. To study the molecular mechanism of low levels of apoB in these individuals, we employed a previously reported knock out mouse model generated by targeted modification of the apob gene. The heterozygous, apoB-100/B-81, mice express full length and truncated apoB, B-81, and have 20 and 35% lower levels of total cholesterol and apoB, respectively, when compared to WT (apoB-100/B-100) mice. The majority of the truncated apoB, B-81, fractionated in the VLDL- density range. The mechanism of low levels of apoB in B-100/B-81 mice was examined. Total hepatic apoB mRNA levels decreased by 15%, primarily due to lower levels of apoB-81 mRNA. Since apoB mRNA transcription rates were similar in B-100/B-100 and B-100/B-81 mice, low levels of mutant apoB-81 mRNA occurred by enhanced degradation of apoB mRNA transcript containing premature translational stop codon. ApoB synthesis measured on isolated hepatocytes decreased in B-100/B-81 mice by 35%, while apoB-48, apoE, and apoAI syntheses remained unchanged. Metabolic studies using whole animal showed a 32% decrease in triglyceride secretion rates, consistent with the apoB secretion rates. Inhibition of receptor-mediated clearance of apoB-81-containing particles resulted in greater relative accumulation of apoB-81 in plasma than apoB-100, suggesting enhanced clearance of apoB-81-containing particles. These results demonstrate that low levels of apoB in heterozygous hypobetalipoproteinemic mice occurs by low rates of apoB secretion, and increased clearance of truncated apoB. Similar mechanisms appear to contribute to low levels of apoB in hypobetalipoproteinemic humans.

Animals↗

PubMed1999

Lipoprotein(a) levels in relation to albumin concentration in childhood nephrotic syndrome.

BACKGROUND: Lipoprotein(a) [Lp(a)] is a lipoprotein consisting of a low-density lipoprotein (LDL) particle linked to a polymorphic glycoprotein, apoprotein(a) [apo(a)]. Prior studies have reported high Lp(a) levels in the nephrotic syndrome, but it is still controversial whether this is due to the degree of hypoalbuminemia or proteinuria. METHODS: To investigate a model of nephrotic syndrome in the absence of renal failure, we studied a group of 84 children in different clinical stages of the disease for a period of five years. We evaluated the direct relationships between lipoproteins, including Lp(a), and/or plasma albumin and proteinuria. RESULTS: Lp(a) levels were significantly higher in the subjects with the active disease compared with patients in remission, and were also significantly different when subjects were ranked by albumin quartiles. Multiple regression analysis revealed that Lp(a) levels were inversely correlated with apo(a) isoform size and plasma albumin levels but not with the proteinuria/creatinine clearance ratio. Among subjects in complete remission, Lp(a) levels were different in patients with albumin levels below or above the fifth percentile. After the improvement of the clinical stage of the disease, the Delta% variation of albumin levels was related to the Delta% of apoB and LDL cholesterol (LDL-C), but not with the Delta% variation of Lp(a), whereas the Delta% variation of LDL-C was, in turn, related to the Delta% of Lp(a) levels. CONCLUSIONS: These results suggest that in the childhood nephrotic syndrome, the increased Lp(a) levels are mainly related to hypoalbuminemia, probably through a mechanism involving apoB overproduction, which leads to an increased number of LDL particles to be converted into Lp(a).

Apolipoproteins B↗

PubMed1999

Molecular bases of low production rates of apolipoprotein B-100 and truncated apoB-82 in a mutant HepG2 cell line generated by targeted modification of the apolipoprotein B gene.

In subjects with familial hypobetalipoproteinemia heterozygous for truncated forms of apolipoprotein B, both apoB-100 and the truncated forms are produced at lower than expected rates. We studied the mechanism of low levels of apoB in a cell model produced by targeted modification of the apob gene of HepG2 cells. One of the three alleles of apob was found to be targeted. The targeted cells expressed apoB-100 and B-82. The media of mutant cells contained 56% of the levels of apoB-100 present in the media of wild-type (WT) HepG2 cells. ApoB-82 was present at 11% of the apoB-100 levels in mutant cell media. An 85-kD protein (apoB-15) representing the N-terminal fragment of apoB was also secreted, but only in the mutant cell media. We examined the mechanism of low levels of apoB-82. Cellular apoB-82 mRNA was 11% of apoB-100 mRNA, lower than the 33% expected, but consistent with relative levels of apoB-82 in the media. ApoB mRNA transcription in WT and the mutant cells did not differ, while the levels of apoB-82 mRNA in nuclei and polysomes were 46% and 12% of the levels of apoB-100 mRNA, respectively, suggesting that the lower levels of apoB-82 mRNA were due to altered message stability. In a pulse/chase experiment with [35S] methionine, at zero time of chase, the amounts of apoB-100 in mutant cells was 66% that of WT levels, consistent with the modification of one allele. The fractions of newly synthesized apoB-100 secreted into the media at 2 h were 10% in the mutant cells and 19% in the WT cells, suggesting greater presecretory degradation of apoB-100 in the mutant cells. Thus, low levels of mutant apoB-82 mRNA gave rise to the low levels of apoB-82, while low levels of apoB-100 were due to low rates of secretion.

Alleles↗

PubMed1999

Known mutations of apoB account for only a small minority of hypobetalipoproteinemia.

Low LDL cholesterol and apoB levels in plasma cosegregate with mutations of apoB in some kindreds with familial hypobetalipoproteinemia. Approximately 35 apoB mutations, many specifying apoB truncations, have been described. Based on the centile nomenclature where the full-length nature apoB consisting of 4536 amino acids is designated as apoB-100, only those truncations of apoB >25% of normal length are detectable in plasma. Previously, we reported on five unrelated kindreds with familial hypobetalipoproteinemia in whom although no apoB truncations were detectable in plasma, low apoB levels were nevertheless linked to the apoB gene. In one of those kindreds, we reported a donor splice site mutation in intron 5 (specifying apoB- 4). We now describe a nonsense mutation in exon 10 (apoB-9) in two of the other unrelated families. Both the apoB-4 and apoB-9 mutations have been reported by others in unrelated families. Recurrent mutations of apoB-40 and apoB-55 also have been reported, suggesting that recurrent mutations of apoB may account for an appreciable proportion of familial hypobetalipoproteinemia kindreds. To test this hypothesis, we searched for four apoB mutations whose products are not detected in plasma including the apoB-4, apoB-9, and two other previously reported mutations in exons 21 and 25. We studied three groups with plasma cholesterols <130 mg/dl in whom no apoB truncations were detected in plasma: a) 28 FHBL probands from St. Louis, b) 151 individual St. Louisians, and c) 28 individual Sicilians. One subject from the 28 kindreds and two subjects among 151 hypobeta individuals from St. Louis harbored the exon 10 mutation. None of the other mutations were detected. Thus, among hypobeta lipoproteinemic subjects without any detectable apoB truncations in plasma, <5% had an apoB truncation-producing mutation. As only about 0.5% of hypobeta lipoproteinemic subjects have plasma-detectable apoB truncations, our data suggest that the known apoB truncations account for only a small proportion of hypocholesterolemia.

Apolipoproteins B↗