Biomedical subjects

Shiu-Ming Kuo

Publications and source records attributed to Shiu-Ming Kuo.

9 recordsLinked to original sources

PubMed2005

Polarized localization of vitamin C transporters, SVCT1 and SVCT2, in epithelial cells.

Messenger RNA of homologous sodium-vitamin C cotransporters, SVCT1 and SVCT2, were found in the intestine. Studies using cultured intestinal cells suggested an apical presence of SVCT1 but the function of SVCT2 was unknown. Here, we showed that enterocytes from heterozygous SVCT2-knockout mice had lower sodium-dependent vitamin C accumulation compared to those from the wildtype. Thus, SVCT2 appears to be functional in enterocytes. We then tested whether SVCT2 could have a redundant function as SVCT1 by constructing and expressing EGFP-tagged SVCTs in intestinal Caco-2 and kidney MDCK cells. In confluent epithelial cells, SVCT1 protein expressed predominantly on the apical membrane. SVCT2, in contrast, accumulated at the basolateral surface. Functionally, SVCT1 expression led to more transport activity from the apical membrane, while SVCT2 expression only increased the uptake under the condition when basolateral membrane was exposed. This differential epithelial membrane distribution and function suggests non-redundant functions of these two isoforms.

Animals↗

PubMed2005

Endotoxin increases ascorbate recycling and concentration in mouse liver.

Sublethal exposure to Escherichia coli endotoxin [lipopolysaccharide (LPS)] attenuates the lethal effects of subsequent insults associated with oxidative stress, such as higher LPS dose, septic peritonitis, and ischemia. Because administration of the antioxidant ascorbate protects against these same insults and injection of dehydroascorbic acid (DHAA) protects against ischemia, the hypothesis that sublethal LPS increases endogenous ascorbate concentration and recycling (i.e., synthesis from DHAA) was tested. Injection of LPS [5 x 10(6) endotoxin units/kg body weight, i.p.] in mice caused a temporary inhibition of food intake, which was significant by 20 h and recovered within 3 d. LPS increased ascorbate concentration in adrenal gland, heart, kidney, and liver. LPS had similar effects in wild-type and Slc23a2+/- mice despite the latter's deficiency in the ascorbate transporter SVCT2. In liver, the ascorbate response to LPS was not accompanied by change in glutathione concentration. LPS decreased gulonolactone oxidase activity, which is rate-limiting for de novo synthesis of ascorbate from glucose, but increased the rate of DHAA reduction to ascorbate. In conclusion, sublethal endotoxin increases ascorbate recycling in liver and ascorbate concentration in liver, adrenal gland, heart, and kidney. The enhanced rate of ascorbate production from DHAA may protect these organs against the reactive oxygen species produced by subsequent, potentially lethal challenges.

Adrenal Glands↗

PubMed2004

Dietary isoflavones suppress endotoxin-induced inflammatory reaction in liver and intestine.

Dietary isoflavone intake has been linked to cancer prevention and their anti-inflammation activity was examined. Intraperitoneal lipopolysaccharide (LPS) injection in mice led to a decrease in the liver antioxidant glutathione level but this decrease was prevented in mice fed with an isoflavone-containing diet. Similarly, isoflavone diet prevented the inflammation-associated induction of metallothionein (MT) in the intestine; and the induction of manganese superoxide dismutase (Mn-SOD) in the liver. Results from the intestinal cell studies suggest that isoflavones suppress the intestinal response to inflammation by modulating the action of pro-inflammatory cytokine, IL-6. IL-6 secretion and the STAT3 (signal transducer and activator of transcription protein 3) nuclear translocation in response to IL-6 were both decreased by genistein.

Animals↗

PubMed2004

Gender and sodium-ascorbate transporter isoforms determine ascorbate concentrations in mice.

We evaluated the hypothesis that sodium-dependent vitamin C (ascorbate) transporters SVCT1 and SVCT2 (encoded by genes Slc23a1 and Slc23a2) regulate ascorbate concentrations in tissues of adult mice. Slc23a2+/- and Slc23a2+/+ mice were fed an ascorbate-free diet for 10-12 wk, and then segregated according to gender and genome, and were placed in groups of 3-4 in metabolic cages for 24-h urine collection. RT-PCR analysis showed that liver and kidney expressed mainly SVCT1, and brain, skeletal muscle, and spleen expressed predominantly SVCT2. Slc23a2+/- mice had low SVCT2 but normal SVCT1 messenger RNA (mRNA) levels, which did not differ between genders. Ascorbate concentrations were lower in Slc23a2+/- than Slc23a2+/+ mice in tissues where SVCT2 was the main isoform. Compared with males, females had lower ascorbate excretion and ascorbate:creatinine ratio in urine and had higher ascorbate concentrations in plasma and SVCT1-predominant tissues. SVCT2 contributed to a gender effect in spleen because males had higher spleen ascorbate concentration than females in wild-type but not in Slc23a2+/- mice. Hepatic gulonolactone oxidase mRNA and activity levels did not differ with genotype or gender, indicating no differences in ascorbate synthesis. We concluded that SVCT2 is a major determinant of ascorbate accumulation in tissues lacking SVCT1. The SVCT isoforms appear to function independently of one another because SVCT1 expression and ascorbate concentrations in SVCT1-predominant organs were not affected by SVCT2 deficiency. Additionally, lower ascorbate excretion in females may elevate the vitamin's concentrations in plasma and tissues expressing SVCT1 that, unlike SVCT2, is not saturated by plasma ascorbate concentrations.

Animals↗

PubMed2004

Cellular vitamin C accumulation in the presence of copper.

Under the cell-free condition, copper is known to oxidize ascorbic acid (the active form of vitamin C) and the event leads to the loss of vitamin C. However, the biological consequence of this interaction was never examined in the presence of cells. We demonstrated in intestinal epithelial cells that dehydroascorbic acid (the oxidized form of ascorbic acid), when generated from ascorbic acid in the presence of copper, can be efficiently transported into the cells and reduced back to ascorbic acid. We also observed in other types of cells the transport and intracellular reduction of dehydroascorbic acid in the presence of copper. In the presence of iron, a metal that also oxidizes ascorbic acid, we observed similar oxidation-related accumulation in intestinal cells. Other metals that do not interact with ascorbic acid had little effect on vitamin C transport. A nonmetal pro-oxidant, hydrogen peroxide, is known to oxidize ascorbic acid and we observed that the oxidation is also accompanied by an increased intracellular accumulation of vitamin C. The efficient coupling between dehydroascorbic acid transport and intracellular reduction could help to preserve the important nutrient when facing oxidative metals in the intestine.

Animals↗

PubMed2003

Estrogen response element-independent regulation of gene expression by genistein in intestinal cells.

Isoflavone, genistein, was shown to have antioxidant and antitumor activities. We have reported the stimulatory effect of genistein on the expression of antioxidant and metal-binding protein, metallothionein IIA (MTIIA), in human intestinal Caco-2 cells. Genistein has been shown to up-regulate the expression of other genes through estrogen response element (ERE) but the ERE sequence is not in the MTIIA promoter. In this paper, we investigated the ERE-independent mechanism that mediates the stimulatory effect of genistein. Genistein enhanced the expression of human MTIIA promoter (up to -426)-containing reporter genes, thus supporting a promoter-specific transcriptional regulation. A shorter MTIIA promoter (-83 to +27) was found to be able to mediate the full reporter gene response to genistein in a dose- and time-dependent fashion. Further deletion and mutation analysis revealed that the GC-rich Sp1 binding sequence was the target of the stimulation. Genistein was known to bind to estrogen receptors (ER). When cells were cotransfected with ER beta, the stimulatory effect of genistein on the reporter gene containing the GC-rich promoter sequence increased further and a similar result was observed for breast cancer MCF-7 cells. Inhibitors of protein kinase A could block the response to genistein but the phosphorylation of Sp1 protein per se was not affected by the genistein treatment. Our observation could help to evaluate the biological significance of genistein, which is used widely as a supplement.

Antineoplastic Agents↗

PubMed2003

Flavonoid structure affects the inhibition of lipid peroxidation in Caco-2 intestinal cells at physiological concentrations.

The antioxidant activity of flavonoids in cell-free systems has been studied extensively. We compared flavonoids with different structural features on their abilities to protect live Caco-2 intestinal cells from lipid peroxidation due to hydrogen peroxide and Fe(2+) treatment. Flavonoids with o-dihydroxyl or vicinal-trihydroxyl groups, including quercetin, myricetin (flavonol), luteolin (flavone) and (-)-epigallocatechin gallate (EGCG; flavanol), when co-incubated with a mixture of 30 micro mol/L H(2)O(2) and 30 micro mol/L FeSO(4), prevented the formation of malondialdehyde (MDA) at 1 or 10 micro mol/L in at least one of two separate experiments. In experiments in which flavonoids were preincubated with cells but removed before the 30 micro mol/L H(2)O(2) and Fe(2+) treatment, quercetin at 0.1 micro mol/L, EGCG at 1 micro mol/L and luteolin at 10 micro mol/L exerted protective effects in at least one of two experiments. Kaempferol (flavonol) and the isoflavones, genistein and daidzein, did not prevent lipid peroxidation at 0.1-10 micro mol/L in either co- or preincubation experiments. None of the flavonoids tested at 0.1-10 micro mol/L increased H(2)O(2) and Fe(2+)-induced lipid peroxidation after co- or preincubation. In summary, these observations support the importance of plant-based food items such as vegetables, fruits and teas in the diet.

Caco-2 Cells↗

PubMed2002

Ascorbic acid status in postmenopausal women with hormone replacement therapy.

OBJECTIVES: Hormone oral contraceptives affected ascorbic acid status adversely in young women. In vitro, estrogens and progesterone inhibited ascorbic acid accumulation in intestinal cells. This is a pilot study to examine the relation between hormone replacement therapy (HRT) and plasma ascorbic acid levels among a group of healthy non-smoking postmenopausal women. METHODS: Healthy non-smoking postmenopausal women aged 48-72 years, 34 with HRT and 21 without HRT, were recruited in summer, 1997. Their fasting plasma ascorbic acid levels were measured and information on ascorbic acid intakes (diet and supplements) was collected through questionnaires. RESULTS: Women taking HRT in this study did not have significantly lower plasma ascorbic acid levels compared with non-HRT users. When subjects were further divided into groups based on ascorbic acid supplementation, HRT users without supplement had a lower mean plasma ascorbic acid level (54+/-16 microM, n=10) compared with non-HRT users (66+/-14 microM, n=12) (P=0.08 for the effect of therapy). HRT users and non-users taking ascorbic acid supplement had similar plasma levels (66+/-10 microM, n=24; 66+/-12 microM, n=9, respectively). CONCLUSION: HRT does not affect ascorbic acid status of healthy well-nourished non-smoking postmenopausal women that are using ascorbic acid supplement. Future larger case-control or supplement intervention study is needed.

Aged↗

PubMed2002

Flavonoids and gene expression in mammalian cells.

Flavonoids appear to regulate the expression of many genes. As expected, when multiple flavonoids were compared in one study, structure-specificity was always observed. Unfortunately, little information is available regarding the proportion of contribution of various structural elements. Also, we have very limited information on their molecular mechanisms of action. The affinity of flavonoids for ER could explain the stimulatory effect on genes with ERE but other modes of action apparently also exist and need to be further explored. Physiological relevance is always a concern when investigating the regulation of gene expression by environmental chemicals such as flavonoids. One factor of concern is the in vivo concentration of flavonoids. Besides intestinal cells, liver cells and skin cells, other tissues obtain flavonoids through blood circulation. Thus, plasma concentrations of flavonoids are normally discussed. Steady state plasma concentrations of flavonoids are usually not much higher than 1 microM even in populations that consume large amounts of plant material. This concentration is relatively low compared to the concentrations of flavonoids that were commonly used in cell culture systems to demonstrate their effectiveness. Nevertheless, we have evidence that some flavonoids may accumulate in the cell. The effect of quercetin on metallothionein expression in Caco-2 cells persisted for at least 24 hours after its removal from the culture medium (Kuo et al., 1998). Also, long-term treatment of cultured cells with quercetin at low concentrations led to a similar effect on metallothionein expression as one high concentration treatment (Kuo et al., 2001). If intracellular accumulation of certain flavonoids is a shared characteristic for various cell types, it implies that routine ingestion of flavonoids could lead to biological effects at the concentration lower than predicted from a single treatment. Experiments to address possible cell/tissue accumulation of flavonoids are greatly needed.

Animals↗