Genetic regulation of cholesterol absorption and plasma plant sterol levels: commonalities and differences

The molecular basis of the processes that control two closely related traits, the absorption of cholesterol from the intestines and plasma plant sterol levels, are only partially understood. The discovery that mutations in two novel hemitransporters, ATP binding cassette transporter G5 (ABCG5) and ABCG8, underlie a rare inborn error in plant sterol metabolism, beta-sitosterolemia, represents a major breakthrough in this field. More recently, genetic studies in the mouse that mapped loci in linkage with cholesterol absorption and plasma plant sterol levels and studies in humans that examined the relationship of plasma plant sterol levels to sequence variation in the ABCG5/ABCG8 locus suggested the involvement of other genes. Moreover, studies in beta-sitosterolemic patients, in ABCG5/ABCG8-targeted animals, and on a newly developed cholesterol absorption inhibitor, ezetimibe, suggest commonalities and differences in the regulation of the two traits. This review summarizes the evidence for genetic control of cholesterol absorption and plasma plant sterol levels, presents the evidence for commonalities and differences between the two traits, and discusses recent developments and future perspectives in this field.     

Loci controlling plasma non-HDL and HDL cholesterol levels in a C57BL /6J x CASA /Rk intercross

Plasma non-HDL and HDL cholesterol levels are predictors of cardiovascular diseases. We carried out a genetic cross between two laboratory inbred mouse strains, C57BL/6J and CASA/Rk, to detect loci that control the plasma levels of non-HDL and HDL cholesterol. With regard to non-HDL cholesterol, chow-fed CASA/Rk males and females had 87% and 25% higher levels, respectively, than did C57BL/6Js. The levels of non-HDL cholesterol in F1s were similar to C57BL/6J. There was no strain difference in HDL cholesterol levels. An intercross between F1s was performed, and plasma non-HDL and HDL cholesterol was measured in 185 male and 184 female mice. In both male and female F2 mice, plasma non-HDL and HDL cholesterol levels were unimodally distributed; however, in both cases the values for females were significantly lower than for males. Therefore, linkage analysis was performed with sex as a covariate. Significant linkage for non-HDL cholesterol was found on chromosome 6 at 49 cM (LOD 5.17), chromosome 4 at 55 cM (LOD 4.22), and chromosome 8 at 7 cM (LOD 3.68). Significant linkage for HDL cholesterol was found on chromosome 9 at 14 cM (LOD 7.52) and chromosome 8 at 76 cM (LOD 4.69). A significant epistatic interaction involving loci on chromosomes 2 and 5 was also observed for non-HDL cholesterol. In summary, linkage analysis in these cross-identified novel loci confirmed previously identified loci in control of plasma non-HDL and HDL cholesterol and disclosed a novel interaction in controlling non-HDL cholesterol levels in the mouse.     

Immunoglobulin gene conversion: synthesizing antibody diversification and DNA repair

Recent developments in the field of antibody (Ab) diversification have rapidly advanced our understanding of the molecular mechanism underlying these events. Key to these developments was the identification of activation-induced cytidine deaminase (AID) as the central regulator of secondary Ab diversification, and the elucidation of its primary function as a DNA deaminase. Incredibly, current literature suggests the existence of a shared pathway, common to all secondary diversification processes, from which the separate outcomes branch outwards at various points. Immunoglobulin gene conversion (IGC) is one of these mechanisms and is used by a number of vertebrate species in both the development of the pre-immune repertoire and in affinity maturation. In a manner similar to other Ab diversification mechanisms, IGC has managed to co-opt a normal DNA repair pathway for the generation of receptor diversity. In the case of IGC specifically, that pathway is homologous recombination (HR). A burgeoning wealth of genetic, biochemical and structural data has clarified the roles of many key HR factors, allowing new insight into its molecular mechanism. These insights, combined with those from the common mechanism of AID action, synergize to develop an emerging picture of the mechanism underlying IGC.     

Using minced horseradish roots and peroxides for the deodorization of swine manure: a pilot scale study

Enzymes that have proven to be capable of removing toxic compounds from water and soil may also be useful in the deodorization of animal manures. Considering that pork production in the US is a $40-billion industry with over half a million workers, odor control to protect air quality in the neighboring communities must be considered an essential part of managing livestock facilities. This pilot scale (20-120 L) study tested the use of minced horseradish (Armoracia rusticana L.) roots (1:10 roots to swine slurry ratio), with calcium peroxide (CaO(2) at 34 mM) or hydrogen peroxide (H(2)O(2) at 68 mM), to deodorize swine slurry taken from a 40,000-gallon storage pit at the Pennsylvania State University's Swine Center. Horseradish is known to contain large amounts of peroxidase, an enzyme that, in the presence of peroxides, can polymerize phenolic odorants and thus reduce the malodor. Twelve compounds commonly associated with malodor (seven volatile fatty acids or VFAs, three phenolic compounds and two indolic compounds) were used as odor indicators. Their concentration in swine slurry before and after treatment was determined by gas chromatography (GC) to assess the deodorization effect. The pilot scale testing demonstrated a complete removal of phenolic odorants (with a detection limit of 0.5 mg L(-1)) from the swine slurry, which was consistent with our previous laboratory experiments using 30-mL swine slurry samples. Horseradish could be recycled (reused) five times while retaining significant reduction in the concentration of phenolic odorants. In view of these findings, inexpensive plant materials, such as horseradish, represent a promising tool for eliminating phenolic odorants from swine slurry.     

High Resistance of Plasmodium falciparum to Sulphadoxine/Pyrimethamine in Northern Tanzania and the Emergence of dhps Resistance Mutation at Codon 581

Background

Sulphadoxine-pyrimethamine (SP) a widely used treatment for uncomplicated malaria and recommended for intermittent preventive treatment of malaria in pregnancy, is being investigated for intermittent preventive treatment of malaria in infants (IPTi). High levels of drug resistance to SP have been reported from north-eastern Tanzania associated with mutations in parasite genes. This study compared the in vivo efficacy of SP in symptomatic 6–59 month children with uncomplicated malaria and in asymptomatic 2–10 month old infants.

Methodology and Principal Findings

An open label single arm (SP) standard 28 day in vivo WHO antimalarial efficacy protocol was used in 6 to 59 months old symptomatic children and a modified protocol used in 2 to 10 months old asymptomatic infants. Enrolment was stopped early (87 in the symptomatic and 25 in the asymptomatic studies) due to the high failure rate. Molecular markers were examined for recrudescence, re-infection and markers of drug resistance and a review of literature of studies looking for the 581G dhps mutation was carried out. In symptomatic children PCR-corrected early treatment failure was 38.8% (95% CI 26.8–50.8) and total failures by day 28 were 82.2% (95% CI 72.5–92.0). There was no significant difference in treatment failures between asymptomatic and symptomatic children. 96% of samples carried parasites with mutations at codons 51, 59 and 108 in the dhfr gene and 63% carried a double mutation at codons 437 and 540. 55% carried a third mutation with the addition of a mutation at codon 581 in the dhps gene. This triple: triple haplotype maybe associated with earlier treatment failure.

Conclusion

In northern Tanzania SP is a failed drug for treatment and its utility for prophylaxis is doubtful. The study found a new combination of parasite mutations that maybe associated with increased and earlier failure.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00361114","term_id":"NCT00361114"}}NCT00361114     

Phytosterolemia on the island of Kosrae: founder effect for a novel ABCG8 mutation results in high carrier rate and increased plasma plant sterol levels

Screening of 932 adults on the Pacific island of Kosrae for plasma plant sterol levels disclosed three subjects, two of them asymptomatic, with phytosterolemia. Sequencing the ATP binding cassette subfamily G member 8 (ABCG8) gene revealed a novel exon 2 mutation that causes a change in codon 24 from glutamine to histidine and a frame shift followed by a premature stop codon, precluding the formation of a functional ABCG8 protein. Genotyping of 1,090 Kosraens revealed 150 as carriers, a 13.8% carrier rate. DNA sequencing of 67 carriers revealed the same mutation as in the probands. In carriers, plasma campesterol and sitosterol levels were 55% and 30% higher, respectively, than in noncarriers. Moreover, compared with noncarriers, carriers showed 21% lower plasma levels of lathosterol, a surrogate marker for cholesterol biosynthesis. There was no difference between the groups in plasma total cholesterol, triglycerides, apolipoprotein B, or apolipoprotein A-I levels. In summary, on the island of Kosrae, a strong founder effect of a mutant ABCG8 allele results in a large number of carriers with increased plasma plant sterol levels and decreased lathosterol levels. The latter finding suggests that heterozygosity for a mutated ABCG8 allele results in a modest increase in dietary cholesterol absorption and a decrease in cholesterol biosynthesis.     

The Effect of Salt Stress on the Production of Canthaxanthin and Astaxanthin by Chlorella zofingiensis Grown Under Limited Light Intensity

The fresh water green microalga Chlorella zofingiensisis known to accumulate ketocarotenoids – primarily astaxanthin but also canthaxanthin – when grown under stress conditions of high light irradiance and low nitrogen. We found that salt stress can replace light stress with respect to inducing carotenoid production: cells of C. zofingiensis grown under low light irradiance and subjected to salt and low nitrogen stress accumulated higher amounts of total secondary carotenoids than those growing under high light and low nitrogen stress. Furthermore, C. zofingiensis growing under conditions of salt stress and low light accumulated higher amounts of canthaxanthin than astaxanthin. It is suggested that for canthaxanthin accumulation under salt stress, light is not a limiting factor, but for astaxanthin accumulation high light irradiance is mandatory. These results may be applied in the future for the commercial production of canthaxanthin by C. zofingiensis in systems in which light availability is poor.     

Hypercholesterolemia and changes in lipid and bile acid metabolism in male and female cyp7A1-deficient mice

Cholesterol 7alpha-hydroxylase, a rate-limiting enzyme for bile acid synthesis, has been implicated in genetic susceptibility to atherosclerosis. The gene, CYP7A1, encoding a protein with this activity, is expressed normally only in hepatocytes and is highly regulated. Our cyp7A1 gene knockout mouse colony, as young adults on a chow diet, is hypercholesterolemic. These mice were characterized extensively to understand how cyp7A1 affects lipid and bile acid homeostasis in different tissue compartments and whether gender plays a modifying role. Both male and female cyp7A1-deficient mice had decreased hepatic LDL receptors, unchanged hepatic cholesterol synthesis, increased intestinal cholesterol synthesis and bile acid transporters, and decreased fecal bile acids but increased fecal sterols. In females, cyp7A1 deficiency also caused changes in hepatic fatty acid metabolism, decreased hepatic canalicular bile acid transporter, Bsep, and gallbladder bile composition altered to a lithogenic profile. Taken together, the data suggest that cyp7A1 deficiency results in a proatherogenic phenotype in both genders and leads to a prolithogenic phenotype in females.