Effect of Ava II and NcoI polymorphisms at the low density lipoprotein receptor gene on plasma lipid levels in a group of Thai subjects.

The contribution of common genetic variations at the LDL receptor gene in determining interindividual differences in plasma lipid levels in the general population has been observed in several studies. In this study, we employed the PCR-RFLP method to investigate such an effect of the common Ava II (exon 13) and Nco I (exon 18) polymorphisms at the low density lipoprotein (LDL) receptor gene locus in 54 normolipidemic Thai subjects. The mean LDL-C level was slightly higher in the Ava II (+/+) genotype than the other Ava II genotypes. This difference was significant at the 5 per cent level although there were only three homozygotes with Ava II (+/+) genotype. The average effect of Ava II (+) allele was to increase LDL-C level by 6.75 mg/dl. A gene-dosage effect was not observed for this polymorphism. In addition, the subjects with Ava II (+/+) genotype also tended to have high serum total cholesterol and triglyceride and low HDL-C levels. Nco I polymorphism revealed no statistically significant effect on lipid levels in these subjects. However, the subjects with (+) allele tended to have high levels of LDL-C, serum total cholesterol and triglyceride.

Screening for a D9N common mutation in exon 2 of the LPL gene in Thai normolipidemic and hyperlipidemic subjects.

Lipoprotein lipase (LPL) is a multifunctional protein, playing a major role in the hydrolysis of triglyceride-rich lipoproteins. It also affects the maturation of high density lipoprotein (HDL) and low density lipoprotein (LDL). A D9N substitution is a frequent mutation found in exon 2 of the LPL gene. It is due to a G --> A transition causing a substitution of Asp by Asn at amino acid residue 9 of the protein. This mutation was screened for in 94 Thai primary dyslipidemic (46 hypercholesterolemic and 48 combined hyperlipidemic) subjects compared to 32 normal healthy subjects using PCR-RFLP. Such a mutation has not, yet, been detected in any of these Thai subjects.

Screening for mutations in exon 4 of the LDL receptor gene in Thai subjects with primary hypercholesterolemia: detection of a novel mutation D151Y by PCR-CFLP.

A mutation in low density lipoprotein (LDL) receptor gene causes an autosomal codominant disorder namely familial hypercholesterolemia (FH). Mutations in the LDL receptor gene are very heterogeneous at the DNA levels, occurring in all 18 exons of the gene. However, exon 4 has been found to be the hot spot for mutational events. In this study DNA from 45 Thai subjects with primary hypercholesterolemia was screened for mutations in the hot spot exon 4. The DNA samples were amplified by Polymerase Chain Reaction (PCR) and screened for mutation by Cleavase Fragment Length Polymorphism (CFLP) technique. Identification of mutation was performed by direct sequencing of PCR product. From this screening, one female patient was found to be heterozygous for a novel mutation which was due to a G to T transversion at nucleotide 514. This transversion would change the species-conserved amino acid at codon 151 from charged R group aspatic (GAC) to uncharged R group tyrosine (TAC), termed D151Y. From the same screening strategy, we found that this mutation was absent in 33 healthy normolipidemic subjects. In this index subject, Arg 3500 Gln mutation in apo B-100 gene, causing hypercholesterolemia namely familial defective apo B-100 (FDB), was not found. Therefore, hypercholesterolemia in this index subject was possibly caused by the D151Y mutation in the LDL receptor gene.

Polymorphism of the gene encoding lipoprotein lipase in thai primary hyperlipoproteinemias.

Lipoprotein lipase (LPL) plays a central role in the clearance of very low density lipoprotein (VLDL) and chylomicrons from the circulation. It also affects the maturation of high density lipoprotein (HDL) and low density lipoprotein (LDL). LPL is an important candidate gene in determining the risk factor in metabolic disorders including primary hyperlipidemia. Our study is the first report from Thailand on the characterization of two common DNA polymorphisms, i.e Pvu II and Hind III at introns 6 and 8, respectively of the LPL gene in 94 Thai dyslipidemic subjects compared to 32 normolipidemic subjects using PCR-RFLP. It was observed that the frequencies of the cut and uncut alleles of Pvu II were 0.67 and 0.33 in normolipidemic subjects. Such frequencies were 0.64 and 0.36 in hyperlipidemic subjects. Additionally, the frequencies of the cut and uncut alleles of Hind III were found to be 0.73 and 0.27 in normolipidemic subjects. They were 0.85 and 0.15 in hyperlipidemic subjects. The allele frequencies of the Hind III but not Pvu II polymorphism in hyperlipidemic subjects were significantly different from normolipidemic subjects (p<0.05). The relation between these polymorphisms and lipid traits was not statistically significant (p>0.05).

Superoxide dismutase and lipid peroxidation in acute lymphocytic leukemia.

There is much evidence suggesting a possible role of reactive oxygen species and other free radicals as mediators of phenotypic and genotypic changes that lead from mutation to cancer. The imbalance in cancer cell antioxidant defense mechanism can influence also the sensitivity to cytoreductive therapy. In erythrocytes, it can result in hemolysis which is one of the pathogenic mechanisms of anemia in cancer patients. Parameters of lipid peroxidation (malondialdehyde, MDA) and antioxidant enzymes here represented by superoxide dismutase (SOD) have been investigated in acute lymphocytic leukemia. Twenty patients of various stages and activities of the disease and twenty five normal controls were recruited. Significantly higher concentrations of total MDA in plasma (28.10 + 2.90 nmol/L vs. 9.20 + 0.30 nmol/L, p < 0.0001) as well as in erythrocytes (3.00 + 0.20 nmol/gHb vs. 1.20 + 0.10 nmol/gHb, p < 0.0001) were found compared to the control group. Significantly lower activity of SOD (1,966 + 110 U/gHb vs. 3,095 + 60 U/gHb, p < 0.0001) was also observed. No correlations were obtained between SOD activity, MDA concentration in both red cells and plasma with blast cell count and hemoglobin concentration. This study suggests that free radicals are associated with an impaired antioxidant defense enzyme activity resulting in increased lipid peroxidation in acute lymphocytic leukemia.

Role of lipid peroxidation, trace elements and anti-oxidant enzymes in chronic renal disease patients.

Plasma Selenium (Se), Zinc (Zn), Copper (Cu) and Aluminium (Al) levels, red blood cell vitamin E and antioxidant enzymes, glutathione peroxidase (GPX) and catalase activity were studied in 54 patients with renal diseases of different levels of kidney dysfunction. Group I (serum creatinine < 2 mg/dl), Group II (serum creatinine 2-4 mg/dl), Group III (serum creatinine 4.1-8 mg/dl), Group IV (serum creatinine 8.1-12 mg/dl) Group V (serum creatinine > 12 mg/dl); thirty two healthy subjects are controls. Plasma Zn (ug/L) and red blood cell vitamin E (ug/ml PRC) were decreased more significantly than controls (1348.59 +/- 43.72 vs 1318.89 +/- 45.62, and 3.38 +/- 0.45 vs 2.23 +/- 0.52) while plasma Selenium and Copper are within normal ranges. Plasma GSH-PX and catalase activity (IU/ml PRC) were also decreased (28.26 +/- 9.01 vs 20.48 +/- 6.79 and 7.54 +/- 1.91 vs 6.52 +/- 2.31) more significantly than controls. Lipid peroxidation products, plasma (umol/L) and urine malonaldehyde (MDA, umol/Ccr) were elevated (7.29 +/- 3.39 vs 92.94 +/- 61.66, and 32.08 +/- 24.60 vs 246.14 +/- 325.66) significantly (p < 0.0001). The lipid peroxidation abnormalities were seen in patients with normal renal function, which supports the role of oxidative stress early in the course of renal disease. Urine ammonia per GFR was also increased as well as urine B2m and NAG. There was no correlation between lipid peroxidation product (MDA) and any of the antioxidant enzymes, vitamin E, urine NH3, B2m, protein or NAG except urine ammonia and MDA per nephron which correlate with severity of kidney dysfunction which confirmed the role of complex processes in the progression of chronic renal failure. The early intervention to decrease oxygen consumption either by dietary protein restriction antioxidants such as vitamin E supplement or calcium channels blockers may be of value in preserving renal function in the setting of chronic renal failure.

Red cell and plasma calcium, copper and zinc in beta-thalassemia/hemoglobin E.

Beta-thalassemia/Hb E is a genetic disease prevalent in Thailand. This study has used atomic absorption spectroscopy to evaluate red cell and plasma calcium, copper and zinc in patients with beta-thalassemia/Hb E, both splenectomized and non-splenectomized. The levels of these trace elements in both red cells and plasma were different between the non-thalassemic controls and the disease patients. The most prominent result was that calcium concentration in red cells increased significantly in thalassemia subjects, particularly in splenectomized cases. These results might reflect the abnormal trace element metabolism and defects in the calcium transport system of the red cell membrane in thalassemia.

The ramification of oral vitamin B-1, C and E on oxidative stress and haematological parameters in normal red blood cells.

Oxidative stress is the state that intimates a causative factor in accelerated cell aging and premature cell death. Our purpose was to study the ability of vitamin B1, C and E in reducing the oxidative stress. In this experiment, we divided the normal subjects into three groups. The first group was supplemented with vitamin B1 (30 mg/day). We found that the superoxide dismutase (SOD) activity and amount of malonyldialdehyde (MDA) reduced significantly when compared with controls (pre-treatment). However, these values increased gradually after supplemented vitamin B1 cessation. The determination of plasma vitamin C and E was also performed and found that vitamin C content increased significantly, but vitamin E level remained constant throughout the experiment. The second group was supplemented with vitamin C (300 mg/day). The enzyme SOD activity, MDA and vitamin E contents were not statistically different during the period of experiment. Nevertheless, the level of plasma vitamin C increased significantly. The last group was supplemented with vitamin E (100 mg/day) with their results equivalent to vitamin B1’s. In addition to these findings, we found that these three vitamins have not effect on hematological data of normal subjects. These results signified that the appropriate antioxidants to prevent oxidant stress were vitamin B1 and E. For vitamin C, it could not reduce oxidative stress, but tended to enhance oxidant stress in the cells.