Effects of statins on the secretion of human serum albumin in cultured HepG2 cells.

Statins reduce cholesterol biosynthesis by inhibiting HMG-CoA reductase and thereby lower total cholesterol and LDL cholesterol levels in serum, which in turn lower the incidence of cardiovascular disease (CVD). Statins are also known to modulate various cellular functions such as gene expression, cell proliferation, and programmed cell death through inhibition of downstream intermediates in cholesterol synthesis. In this study, we have investigated the possible effects of statins on the secretion of serum albumin from cultured HepG2 cells since high levels of serum albumin are associated with reduced risks for CVD and statins are effective in lowering the risk of CVD through other effects in addition to their effects on serum total cholesterol and LDL cholesterol levels, known as pleiotropic effects. Our results showed that simvastatin increased HSA secretion up to 32.3% compared to the control group. Among 3 statin analogs we tested, simvastatin exhibited the highest stimulatory effects on HSA secretion compared to the control group. Our study also showed that the increased HSA secretions from HepG2 cells by simvastatin treatments were due to the increased rate of HSA synthesis, not due to the reduced posttranslational degradation rate of HSA. Our finding suggests another added benefit of statins' treatments in preventing CVD through stimulation of HSA biosynthesis.

Association of genetic variation in the transforming growth factor beta-1 gene with serum levels and risk of colorectal neoplasia.

In the normal intestinal epithelium transforming growth factor beta-1 (TGFbeta-1) acts as a growth inhibitor, but in malignant cells it may act as a tumor promoter. However, only limited information is available on genetic variation in the TGFB1 gene and its relationship to circulating levels and risk of colorectal cancer. To characterize associations of genetic variation [tagging single-nucleotide polymorphisms (tagSNP) and haplotypes with frequency >0.05] at the TGFB1 locus with circulating TGFbeta-1 and risk of colorectal neoplasia, we conducted two case-control studies (including 271 colorectal adenoma cases and 544 controls, and 535 colorectal adenocarcinoma cases and 656 controls) among Japanese Americans, Caucasians, and Native Hawaiians in Hawaii. Serum TGFbeta-1 was measured by sandwich ELISA among the subjects of the first study. The variant A allele for tagSNP rs6957 was associated with higher serum TGFbeta-1 [means (in ng/mL) and 95% confidence interval (95% CI) for AA or AG, 32.6 (30.6-34.7); GG, 29.0 (25.1-32.9); P(difference) = 0.05] after adjusting for age and other factors. Homozygous carriers of the variant G allele for tagSNP rs11466345 had a statistically significantly lower risk of adenocarcinoma [AG versus AA: odds ratio (OR), 0.9 (95% CI, 0.7-1.2); GG versus AA: OR, 0.4 (95% CI, 0.2-0.7); P(trend) = 0.01]. The haplotype carrying both variants was also statistically significantly associated with a reduced risk of adenocarcinoma (OR, 0.3; 95% CI, 0.1-0.8). Although not statistically significant, the direction and magnitude of the corresponding ORs were similar for adenoma. These results suggest that a haplotype containing SNP rs11466345 at the 3' end of TGFB1 is associated with genetic susceptibility to colorectal neoplasia.

Cruciferous indole-3-carbinol inhibits apolipoprotein B secretion in HepG2 cells.

The cardioprotective effect of consuming cruciferous vegetables may be attributed to a number of unique indole-based compounds. We investigated the potential role and mechanism of action of an indole-based compound, indole-3-carbinol (I-3-C), on apolipoprotein B-100 (apoB) production using HepG2 cells. I-3-C reduced apoB secretion into the media dose dependently by 56% at 100 micromol/L. Relative to the untreated control cells, no change in the density of the secreted lipoproteins was noted. Significant decreases in cellular lipid synthesis, including triglycerides (TG) and cholesterol esters (CE), were observed in cells treated with I-3-C, indicating that limited lipid availability is a major factor in the regulation of apoB secretion. The decrease in TG synthesis was associated with significantly decreased diacylglycerol acyltransferase-1 and -2 activity and reduced fatty acid synthase (FASN) gene expression. The decreased CE synthesis was associated with significantly decreased acyl CoA:cholesterol acyltransferase gene expression and activity. The effect on FASN was shown to be mediated by sterol regulatory element binding protein-1, an important transcription factor involved in fatty acid synthesis. Further investigative work revealed that LDL uptake and fatty acid oxidation were not involved in the I-3-C-mediated reduction of apoB secretion. The results indicate that plant indoles have beneficial effects on lipid synthesis that could contribute to their potential cardioprotective effect.

Impaired glucose tolerance is associated with postganglionic sudomotor impairment.

We compared quantitative sudomotor axon-reflex test responses in persons with normal and impaired glucose tolerance (IGT). Responses were significantly impaired in those with IGT, which may be indicative of early distal small fiber neuropathy.

Prevalence of diabetes and glucose intolerance in an ethnically diverse rural community of Hawaii.

BACKGROUND: We report the prevalence of diabetes in a rural, multiethnic community in Hawaii, of predominantly Asian and Native Hawaiian ancestry, by using 1997 World Health Organization diagnostic criteria applied to a two-hour oral glucose tolerance test. METHODS: This cross-sectional survey included 1452 men and nonpregnant women who were >18 years of age. Blood was drawn in the fasting and postchallenge states. Individuals under pharmacologic treatment for diabetes were excluded. Information obtained included demographics, medical history, dietary intake, physical activity, and anthropometric measurements. RESULTS: Prevalence of diabetes was approximately three-fold higher among Asian and Native Hawaiian ancestry groups than among Caucasians, even after adjusting for other risk factors. Furthermore, diabetes prevalence was similar among all non-Caucasian ethnic groups despite significant differences in body mass indices. CONCLUSIONS: These findings indicate that earlier reports of high prevalence of diagnosed diabetes among Asians and Hawaiian ethnic groups were not due to detection bias, since our study revealed similar prevalence of previously unrecognized diabetes. Furthermore, similar prevalence among these groups was observed despite significant differences in body mass indices, diet, and physical activity. This apparent paradox may reflect limitations in the measurement of these risk factors; differences in the impact of these risk factors on diabetes risk in different ethnic groups; or ethnic differences in lifestyle, biochemical, or genetic factors that were not examined in this study.

Association between angiotensin-converting enzyme gene polymorphisms and QT duration in a multiethnic population in Hawaii.

OBJECTIVES: Recent studies have suggested that heart-rate corrected QT interval (QTc) in normal populations may be influenced by genetic factors. We report findings of a study of the relationship between QTc, increased QTc (> 440 ms) and angiotensin-converting enzyme (ACE) genotype in a multiethnic, population-based study completed in rural Hawaii. METHODS: Blood samples were obtained while fasting and after an oral glucose challenge from 1452 individuals between 1997 and 2000. The clinical examination included an electrocardiogram. Medical histories, behavioral and socio-demographic information were obtained during the interview. Ethnicity was estimated by self-report. The insertion/deletion (I/D) polymorphism in intron 16 of the ACE gene was determined by polymerase chain reaction (PCR) from a random sample of 588 participants. Multiple linear and logistic regression was used to test for associations between QTc and ACE gene polymorphisms. RESULTS: The overall crude prevalence of increased QTc was 21.2%. The prevalence of increased QTc was lowest among those with ACE DD genotype, and highest among those with ACE insertion/insertion (II) genotype. The adjusted odds ratio for increased QTc was 2.29 (95% CI 1.02-5.12) and 3.61 (95% CI 1.60-8.13) for ID and II genotypes, respectively, compared to the DD genotype. The test for trend was highly significant (p < 0.001). CONCLUSIONS: The ACE insertion allele was associated with increased prevalence of prolonged QTc independent of ethnicity, age, gender, and BMI. These findings may implicate the ACE gene as an important genetic risk factor for cardiovascular disease morbidity and mortality.

Citrus polymethoxylated flavones improve lipid and glucose homeostasis and modulate adipocytokines in fructose-induced insulin resistant hamsters.

The present study was undertaken to determine whether supplementation with polymethoxylated flavones (PMFs) could ameliorate the fructose-induced hypertriglyceridemia and other metabolic abnormalities associated with insulin resistance (IR) in hamsters. Following feeding with the fructose diet, hamsters were supplemented orally with PMF-L or PMF-H (62.5 and 125 mg/kg/day) for 4 weeks. Both PMF-treated groups showed a statistically significant (p<0.05) decrease in serum triglyceride (TG) and cholesterol levels compared to the fructose-fed control group. The fructose control group at the end of the study showed elevated serum insulin and impaired insulin sensitivity (glucose intolerance). On the other hand, PMF-supplemented groups showed a reversal in these metabolic defects, including a decrease in insulin level and an improvement in glucose tolerance. PMF supplementation also reduced TG contents in the liver and heart and was able to regulate adipocytokines by significantly suppressing TNF-alpha, INF-gamma, IL-1beta and IL-6 expression and increasing adiponectin in IR hamsters. The mechanism of PMF on the activation of peroxisome proliferator-activated receptors (PPAR) was also explored. PMF-H supplementation significantly increased PPARalpha and PPARgamma protein expression in the liver. This is the first report of positive effects of PMF on adipocytokine production and on PPAR expression in IR hamsters. This study suggests that PMF can ameliorate hypertriglyceridemia and its anti-diabetic effects may occur as a consequence of adipocytokine regulation and PPARalpha and PPARgamma activation.

Green tea leaf extract improves lipid and glucose homeostasis in a fructose-fed insulin-resistant hamster model.

The present study evaluated the effect of green tea (Camellia sinensis L.) leaf extract on triglyceride and glucose homeostasis in a fructose-fed hypertriglyceridemic, insulin-resistant hamster model. There was a significant decrease in plasma triglyceride levels following supplementation of the green tea epigallocatechin gallate-enriched extract (42% at 150 mg/(kg day) to 62% at 300 mg/(kg day) for 4 weeks). Compared to baseline, the fructose control group at the end of the study showed elevated serum insulin and apolipoprotein B levels, and decreased serum adiponectin levels. The fructose/green tea extract group showed a reversal in all of these metabolic defects, including an improvement in glucose levels during a glucose tolerance test. Triglyceride content was also examined in various tissues and compared to the control fructose group; supplementation of the green tea extract (300 mg/kg) reduced triglyceride content in liver and heart tissues. There was molecular evidence of improved lipid and glucose homeostasis based on peroxisome proliferator-activated receptor (PPAR) protein expression. Compared to the control fructose group, supplementation of the green tea extract (300 mg/kg) significantly increased PPARalpha and PPARgamma protein expression. In summary, the data suggest that intake of the green tea extract ameliorated the fructose-induced hypertriglyceridemia and the insulin-resistant state in part through PPAR.

Increased diacylglycerol acyltransferase activity is associated with triglyceride accumulation in tissues of diet-induced insulin-resistant hyperlipidemic hamsters.

Over-accumulation of triglyceride (TG) in insulin-sensitive tissues is associated with the development of insulin resistance. We investigated whether enhanced de novo lipogenesis via diacylglycerol acyltransferase (DGAT) may contribute to the over-accumulation of TG in various tissues (liver, adipose, muscle, and intestine) using 2 well-characterized hyperlipidemic, insulin-resistant hamster models. In general, a marked increase in TG accumulation was noted in most tissues. Interestingly, the increase in TG accumulation corresponded to an increase in microsomal DGAT activity which ranged from 114% to 575% in all of the examined tissues (n = 7 per group). To delineate the mechanism for the increase in DGAT activity, we measured the expression of DGAT-1 and DGAT-2 messenger RNA by relative reverse transcriptase polymerase chain reaction (RT-PCR). In general, DGAT gene expression changed with DGAT-1 changing the most in the liver and adipose tissue, whereas DGAT-2 showed responses mainly in muscle and intestine. The increases in messenger RNA expression were not remarkable (averaging 35%; n = 4 per group) indicating that posttranscriptional mechanism(s) may play a larger role in regulating DGAT activity. In summary, the data suggest that elevated DGAT activity/expression and the subsequent increase in de novo lipogenesis could in part induce the insulin-resistant state.

Microsomal triglyceride transfer protein gene expression and ApoB secretion are inhibited by bitter melon in HepG2 cells.

Momordica charantia or bitter melon is traditionally used as an antidiabetic agent in Asia, Africa, and South America. Recent studies indicate that bitter melon can also lower plasma lipids and VLDL in diabetic animal models as well as animals fed a high-fat diet, suggesting an effect on lipoprotein metabolism. The aim of this study was to delineate the cellular and molecular mechanisms involved in the lipid-lowering properties of bitter melon and regulation of apolipoprotein B (apoB). Human hepatoma cells, HepG2, treated with bitter melon juice (BMJ) for 24 h reduced apoB secretion with and without the addition of lipids (P < 0.05). However, BMJ did not increase apoB secretion in cells treated with N-acetyl-leucyl-leucyl-norleucinal, indicating a lack of effect on the proteasomal degradation pathway. BMJ reduced the secretion of new triglycerides (P < 0.05) and decreased microsomal triglyceride transfer protein (MTP) mRNA expression, suggesting that lipid bioavailability and lipidation of lipoprotein assembly are likely involved in decreased apoB secretion. Interestingly, BMJ increased the nuclear translocation of the mature form of sterol regulatory element-binding protein-1c (SREBP-1c, P < 0.05), involved in MTP secretion. Our data suggest that BMJ is a potent inhibitor of apoB secretion and TG synthesis and secretion that may be involved in the plasma lipid- and VLDL-lowering effects observed in animal studies.

Prevalence and risk factors for prolonged QTc in a multiethnic cohort in rural Hawaii.

BACKGROUND: Few studies have examined the biochemical risk factors for prolonged QTc, a predictor of mortality in numerous studies. We report on the prevalence and risk factors for prolonged QTc in a multiethnic population in rural Hawaii. METHODS: Electrocardiograms were collected from 1415 participants in a cross-sectional survey. The QT interval lengths were corrected for heart rate using Bazett's formula. Linear and logistic regression models were used to examine associations between various cardiovascular risk factors with QTc. RESULTS: Among the CVD risk factors examined, only age, gender, 2-h glucose, and systolic blood pressure (SBP) were independently associated with QTc interval length. Significant ethnic differences in prevalence were also observed, which persisted after controlling for other risk factors. CONCLUSIONS: Significant associations between prolonged QTc and ethnic ancestry, but not cholesterol or triglyceride levels, suggest that genetic factors may play a more important role in determining QTc interval length than conventional biochemical and metabolic CVD risk factors.

Inhibitory activity of diacylglycerol acyltransferase (DGAT) and microsomal triglyceride transfer protein (MTP) by the flavonoid, taxifolin, in HepG2 cells: potential role in the regulation of apolipoprotein B secretion.

The purpose of the present study was to examine the role of taxifolin, a plant flavonoid, on several aspects involving apolipoprotein B (apoB) secretion and triglyceride (TG) availability in HepG2 cells. Taxifolin was shown by ELISA to markedly reduce apoB secretion under basal and lipid-rich conditions up to 63% at 200 micromol/L. As to the mechanism underlying this effect, we examined whether taxifolin exerted its effect by limiting TG availability in the microsomal lumen essential for lipoprotein assembly. Taxifolin was shown to inhibit microsomal TG synthesis by 37% and its subsequent transfer into the lumen (-26%). The reduction in synthesis was due to a decrease in diacylglycerol acyltransferase (DGAT) activity (-35%). The effect on DGAT activity was found to be non-competitive and non-transcriptional in nature. Both DGAT-1 and DGAT-2 mRNA expression remained essentially unchanged suggesting the point of regulation may be at the post-transcriptional level. Evidence is accumulating that microsomal triglyceride transfer protein (MTP) is also involved in determining the amount of lumenal TG available for lipoprotein assembly and secretion. Taxifolin was shown to inhibit this enzyme by 41%. Whether the reduction in TG accumulation in the microsomal lumen is predominantly due to DGAT and/or MTP activity remains to be addressed. In summary, taxifolin reduced apoB secretion by limiting TG availability via DGAT and MTP activity.

The chalcone xanthohumol inhibits triglyceride and apolipoprotein B secretion in HepG2 cells.

The present study examined the role of xanthohumol (XN), a plant chalcone, on apolipoprotein B (apoB) and triglyceride (TG) synthesis and secretion, using HepG2 cells as the model system. The data indicated that XN decreased apoB secretion in a dose-dependent manner under both basal and lipid-rich conditions (as much as 43% at 15 micromol/L). This decrease was associated with increased cellular apoB degradation. To determine the mechanism underlying this effect, we examined triglyceride availability, a major factor in the regulation of apoB secretion. XN inhibited the synthesis of TG in the microsomal membrane and the transfer of this newly synthesized TG to the microsomal lumen (decreases of 26 and 64%, respectively, under lipid-rich conditions), indicating that TG availability is a determining factor in the regulation of apoB secretion under the experimental conditions. The inhibition of TG synthesis was caused by a reduction in diacylglycerol acyltransferase (DGAT) activity, which corresponded to a decrease in DGAT-1 mRNA expression, but not DGAT-2 expression. Microsomal triglyceride transfer protein (MTP) may also control the rate of TG transfer from the microsomal membrane to the active lumenal pool. XN decreased MTP activity in a dose-dependent manner (as much as 30%). Whether the reduction in TG accumulation in the microsomal lumen is predominantly due to DGAT and/or MTP activity remains unknown. In summary, the data suggest that xanthohumol is a potent inhibitor of apoB secretion.

Modulation of HepG2 cell net apolipoprotein B secretion by the citrus polymethoxyflavone, tangeretin.

The purpose of the present study was to examine the role of tangeretin, a polymethoxylated flavone from citrus fruits, on the regulation of apolipoprotein B (apoB) and lipid metabolism in the human hepatoma cell-line HepG2. The marked reduction in apoB secretion observed in cells incubated with 72.8 microM tangeretin was rapid, apoB-specific, and partly reversible. The reduction also was observed under lipid-rich conditions and found to be insensitive to proteasomal degradation of nascent apoB. We followed our study by examining lipid synthesis and mass. A 24-h exposure of cells to 72.8 microM tangeretin decreased intracellular synthesis of cholesteryl esters, free cholesterol, and TAG by 82, 45, and 64%, respectively; tangeretin also reduced the mass of cellular TAG by 37%. The tangeretin-induced suppression of TAG synthesis and mass were associated with decreased activities of DAG acyltransferase (up to -39.0 +/- 3.0% vs. control) and microsomal triglyceride transfer protein (up to -35.5 +/- 2.5% vs. control). Tangeretin was also found to activate the peroxisome proliferator-activated receptor, a transcription factor with a positive regulatory impact on FA oxidation and TAG availability (up to 36% increase vs. control). The data suggest that tangeretin modulates apoB-containing lipoprotein metabolism through multiple mechanisms.