Sureerut Porntadavity

Transcription regulation of human manganese superoxide dismutase gene.

The human MnSOD gene has a typical housekeeping gene promoter, but is highly inducible by various physical, chemical, and biological agents. Transcription factors SP-1 and AP-2 seem to have opposite roles in the transcriptional activity of the basal promoter. Whereas SP-1 plays a positive role, which is absolutely essential for transcription from the human MnSOD promoter, AP-2 appears to play a negative role in this process. An enhancer element is found in the promoter region of the human MnSOD gene. Several important enhancer elements are located in the second intron. The NF-kappa B site in the second intron is essential but not sufficient for high-level induction of MnSOD by cytokines. Although mutations in the regulatory elements may be partially responsible for the lack of induction of MnSOD in some cell types, differences in the degree of induction exist that cannot be accounted for by the defect in the DNA sequence. It is highly likely that this difference is due to the presence or absence of coactivator or suppressor proteins in the cells and may have a physiological role in the defense against oxidative stress.

Effect of atorvastatin on paraoxonase1 (PON1) and oxidative status

It has been proposed that paraoxonase1 (PON1), a high density lipoprotein (HDL)-associated esterase/lactonase, has anti-atherosclerotic properties. The activity of PON1 is influenced by PON1 polymorphisms. However, the influence of PON1 polymorphisms on PON1 activity and oxidative stress in response to lipid-lowering drugs remains poorly understood. The objective of the present study was to investigate the effects of atorvastatin on PON1 activity and oxidative status. The influence of PON1 polymorphisms on PON1 activity and oxidative status in response to atorvastatin treatment was also evaluated. In total, 22 hypercholesterolemic patients were treated with atorvastatin at a dose of 10 mg/day for 3 months. Lipid profile, lipid oxidation markers (malondialdehyde (MDA), conjugated diene (CD), total peroxides (TP)), total antioxidant substance (TAS), oxidative stress index (OSI), and paraoxonase1 activity were determined before and after treatment. L55M, Q192R, and T(-107)C PON1 polymorphisms were also determined. Atorvastatin treatment significantly reduced the levels of total cholesterol (24.5%), low density lipoprotein (LDL) cholesterol (25.4%), triglycerides (24.4%), CD (4.4%), MDA (15.2%), TP (13.0%) and OSI (24.0%), and significantly increased the levels of TAS (27.3%), and PON1 activity (14.0%). Interestingly, the increase in PON1 activity and the reduction in oxidative stress in response to atorvastatin were influenced only by the PON1 T-107C polymorphism. Atorvastatin treatment improved the lipid profile, lipid oxidation, and oxidative/antioxidative status markers including the activity of PON1 towards paraoxon. These beneficial effects may be attributed to the antioxidant properties of statins and the increase in PON1 activity. The increase in PON1 activity was enhanced by the PON1 T-107C polymorphism.

TPA-Activated Transcription of the Human MnSOD Gene: Role of Transcription Factors SP-1 and Egr-1

Induction of manganese superoxide dismutase (MnSOD) in response to oxidative stress has been well established in animals, tissues, and cell culture. However, the role of the human MnSOD (hMnSOD) promoter in stimulus-dependent activation of transcription is unknown. The hMnSOD promoter lacks both a TATA and a CAAT box but possesses several GC motifs. In a previous study, we showed that the basal promoter contains multiple Sp1 and AP-2 binding sites and that Sp1 is essential for the constitutive expression of the hMnSOD gene. In this study, we identified an Egr-1 binding site in the basal promoter of hMnSOD. We also found that the basal promoter is responsive to 12-O-tetradecanoylphorbol-13-acetate (TPA)-activated hMnSOD transcription in the human hepatocarcinoma cell line HepG2. The contributions of these binding sites and the roles of the transcription factors Egr-1, AP-2, and Sp1 in the activation of hMnSOD transcription by TPA were investigated by site-directed mutation analysis, Western blotting, and overexpression of transcription factors. The results showed that Sp1 plays a positive role for both basal and TPA-activated hMnSOD transcription, whereas overexpression of Egr-1 has a negative role in the basal promoter activity without any effect on TPA-mediated activation of hMnSOD transcription.

Decreased paraoxonase 1 activity and increased oxidative stress in low lead−exposed workers

Paraoxonase 1 (PON1) has been proposed as an antioxidant enzyme. Although lead-inhibited PON1 activity has been demonstrated mostly based on in vitro experiments, it is uncertain whether this phenomenon is relevant in pathogenesis of lead-induced oxidative stress in the lead exposure. We examined associations of blood lead levels (BLL) and PON1 activity along with oxidative stress parameters in lead exposure workers. We determined malondialdehyde (MDA), conjugated diene (CD), total peroxides (TP), total antioxidant status (TAS), the oxidative stress index (OSI), and PON1 activity in earthenware factory workers (n = 60) and control subjects (n = 65). The lead-exposed group significantly increased lipid peroxidation parameters and OSI compared to the control group (p < 0.001). The lead-exposed group had significantly decreased PON1 activity and TAS levels compared to the control group (p < 0.001). Multiple linear regression analysis revealed that BLL were significantly correlated with decreased TAS (r = −0.496) and PON1 activity (r = −0.434), but with increased CD (r = 0.694), TP (r = 0.614), MDA (r = 0.788), and OSI (r = 0.722). Interestingly, BLL at 10 µg/dL significantly decreased PON1 activity and increased oxidative stress parameters with insignificant changes in other biochemical and hematological parameters. Altogether, the reduction of PON1 activity may associate in an imbalance in pro-oxidants and antioxidants, leading to oxidative damage in lead-exposed workers even at low BLL.

Effects of Curcuma comosa on the expression of atherosclerosis-related cytokine genes in rabbits fed a high-cholesterol diet

AIM OF THE STUDY Curcuma comosa has been known to have potential use in cardiovascular diseases, but its immunoregulatory role in atherosclerosis development and liver toxicity has not been well studied. We therefore investigated the effects of Curcuma comosa on the expression of atherosclerosis-related cytokine genes in rabbits fed a high-cholesterol diet. MATERIALS AND METHODS Twelve male New Zealand White rabbits were treated with 1.0% cholesterol for one month and were subsequently treated with 0.5% cholesterol either alone, or in combination with 5mg/kg/day of simvastatin or with 400mg/kg/day of Curcuma comosa powder for three months. The expression of IL-1, MCP-1, TNF-α, IL-10, and TGF-β in the isolated abdominal aorta and liver were determined by real-time RT-PCR. Liver toxicity was determined by hepatic enzyme activity. RESULTS Curcuma comosa significantly decreased the expression of pro-inflammatory cytokines, leading to a stronger reduction in IL-1, MCP-1, and TNF-α expression compared to that was suppressed by simvastatin treatment. However, neither Curcuma comosa nor simvastatin affected the expression of anti-inflammation cytokines. In the liver, Curcuma comosa insignificantly decreased the expression of pro-inflammatory cytokines and significantly increased the expression of the anti-inflammatory cytokine IL-10 without altering the activity of hepatic enzymes. In contrast, simvastatin significantly increased the MCP-1 and TNF-α expressions and serum ALT level, without affecting the expression of anti-inflammatory cytokines. CONCLUSIONS In this study, we demonstrated that Curcuma comosa exerts anti-inflammatory activity in the aorta and liver without causing liver toxicity, indicating that Curcuma comosa is a potential candidate as an alternative agent in cardiovascular disease therapy.

Combined PCSK9 and APOE Polymorphisms are Genetic Risk Factors Associated with Elevated Plasma Lipid Levels in a Thai Population

Proprotein convertase subtilisin/kexin type 9 (PCSK9) and apolipoprotein E (ApoE) play a key role in the regulation of lipid metabolism. We aimed to investigate the effects of PCSK9 (R46L, I474V, and E670G) and APOE polymorphisms on lipid levels in a Southern Thai population. A total of 495 participants (307 urban, 188 rural) were recruited for the study. Anthropometric and biochemical variables were evaluated. PCSK9 and APOE polymorphisms were analyzed using PCR–RFLP. The 46L urban male carriers had significantly higher diastolic blood pressure (DBP) and fasting blood sugar compared with non-carriers. In contrast, the 46L urban female carriers had significantly lower total cholesterol (TC) and LDL-C levels compared with non-carriers. The 474V rural female carriers had significantly lower HDL-C levels than non-carriers. The 670G urban female carriers showed significantly higher TC and LDL-C levels compared with non-carriers. APOE4 carriers had increased TC and LDL-C levels relative to APOE3 carriers in the urban males. APOE2 carriers had decreased TC and/or LDL-C levels compared with APOE3 carriers in urban males and females. A significant trend of increased TC and LDL-C levels was observed in non-APOE4-PCSK9 670EE carriers to APOE4-PCSK9 670EG carriers in urban subjects. In summary, R46L, I474V, and E670G may be genetic risk factors for cardiovascular disease (CVD) in urban males, rural females, and urban females, respectively. In contrast, R46L had a favorable lipid profiles that may protect against CVD in urban females. The combination of PCSK9 E670G and APOE polymorphisms may represent an independent factor for the determination of lipid levels.

Lead inhibits paraoxonase 2 but not paraoxonase 1 activity in human hepatoma HepG2 cells

Lead is an environmental toxicant of great concern for humans and animals. Lead‐induced liver damage and malfunction are partly due to a disturbance of the cellular antioxidant balance. Paraoxonase 1 (PON1) and PON2 are highly expressed in the liver and have been proposed as antioxidative enzymes. In this study, the effects of lead on PON1 and PON2 activities were investigated in human hepatoma HepG2 cells by exposing the cells to various concentrations of lead acetate for 24, 48, or 72 h. The results show that a significant increase in reactive oxygen species was observed even at the lowest concentration of lead treatment. However, only the highest concentration of lead significantly influenced cell viability. Lead had no influence on cell‐associated PON1 activity, but it significantly decreased cytoplasmic PON2 activity in a concentration‐ and time‐dependent manner. This reduction was rescued by the addition of calcium. A significant increase of PON2 transcript was observed by real‐time polymerase chain reaction, while PON2 protein expression did not change in the western blot analysis. Taken together, these results indicate that lead reduces PON2, but not PON1, activity and that this reduction is reversed by calcium. Lead‐induced oxidative stress and decreased PON2 activity lead to the upregulation of PON2 transcript. Copyright

Human paraoxonase 2

Human paraoxonase 2 (PON2), which is a member of the paraoxonase family, possesses unique properties that distinguish it from PON1 and PON3. PON2 is ubiquitously expressed in many different tissue types and is highly expressed in the vital organs, such as the heart and lungs. Early research revealed that PON2 is exclusively intracellularly found, wherein it functions as an anti-oxidative protein by reducing intracellular and local oxidative stress. Studies in the last five years have demonstrated that PON2 protects against atherosclerosis by preventing low-density lipoprotein (LDL) oxidation, reversing the oxidation of mildly oxidised LDL, inhibiting monocyte chemotaxis, and increasing cholesterol efflux. Recently, emerging evidence has proposed that PON2 is an anti-atherosclerotic and may be associated with cardiovascular disease (CVD). The number of investigations concerning the relationship between two common PON2 polymorphisms and CVD among different ethnic groups and regions is rapidly growing. Here, we briefly review the developments in PON2 research by focusing on past and recent findings.

Two novel D151Y and M391T LDLR mutations causing LDLR transport defects in Thai patients with familial hypercholesterolemia.

BACKGROUND Familial hypercholesterolemia (FH) is an autosomal dominant disorder caused by mutations in the low density lipoprotein receptor (LDLR) gene. Two novel LDLR mutations, D151Y and M391T, had been previously identified in unrelated Thai patients with heterozygous FH. To confirm that these mutations cause FH, the functional characteristics of D151Y and M391T, which are located in the fourth cysteine repeat of the ligand-binding domain and in the sixth YWTD repeat of the epidermal growth factor precursor homology domain, respectively, were studied. METHODS CHO-ldlA7 cells were transfected with wild type and mutant LDLR cDNAs. Thereafter, the localization, expression, and ability of LDL uptake of LDLR were evaluated by confocal laser scanning microscope (CLSM), and flow cytometry. RESULTS CLSM revealed both D151Y and M391T LDLR were partially retained in the endoplasmic reticulum, with the remaining residual activity observed by LDL uptake. Similarly, flow cytometric analysis showed a significant reduction of LDLR expression to 18% and 38% and of LDL uptake to 15% and 71% in D151Y and M391T LDLR, respectively. CONCLUSIONS The transport defect of LDLR contributes to the pathology of FH. These data are useful for an insight inspires the development of novel lipid-lowering drugs with beneficial therapeutic value.

Molecular modeling of D151Y and M391T mutations in the LDL receptor

The low-density lipoprotein receptor (LDLR) is a key regulator of cholesterol homeostasis, and defects in the function of LDLR result in familial hypercholesterolemia (FH). In the present study, we performed structural analyses of two novel LDLR mutations, D151Y and M391T. Both mutations occurred in conserved residues of LDLR. The D151Y mutation, in the ligand binding domain, caused an elimination of a hydrogen bond in the calcium binding site, higher solvent accessibility and a loss of negative charge in the Y151 residue. On the other hand, the M391T mutation, in the beta-propeller of the epidermal growth factor (EGF) precursor homology domain, caused an additional hydrogen bond to form, higher solvent accessibility and a distortion of the beta-strand. These data suggest that the irregular structures of the mutated LDLRs are likely to cause the functional defect that contributes to the pathology of FH.