Humic acid induces the expression of ox-LDL receptor in HL-60 cells through activation of PPAR-gamma.
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DNA-Binding Proteins↗
Biomedical subjects
F J Lu
Publications and source records attributed to F J Lu.
In vitro anti-influenza virus activity of synthetic humate analogues derived from protocatechuic acid.
Two humic-like substances, the oxidative polymer of protocatechuic acid (OP-PCA) and humic acid inhibit the in vitro replication of influenza virus A/WSN/33 (H1N1) in Madin-Darby canine kidney (MDCK) cells at concentrations of no cytotoxicity. The 50% inhibitory concentration (IC50) for OP-PCA was 6.59 +/- 1.02 microg/ml when the compound was added at the stage of viral adsorption. When OP-PCA was added after virus adsorption, the IC50 was 53.27 +/- 8.12 microg/ml. The IC50 for humic acid was 48.61 +/- 7.32 microg/ml and 55.27 +/- 5.46 microg/ml respectively when the compound was added at the stage of viral adsorption or post-adsorption. In spite of structural resemblance of these two compounds, they exhibit different actions of anti-flu. The OP-PCA inhibits virus-induced hemagglutination and low pH-induced cell-cell fusion. Humic acid inhibits the endonuclease activity of viral RNA polymerase. The monomer of PCA shows no inhibition on influenza virus replication.
Animals↗
Leukocyte mitochondria depolarization and apoptosis in advanced heart failure: clinical correlations and effect of therapy.
OBJECTIVES: The purpose of this study was to examine the changes in leukocyte mitochondrial transmembrane potential (MTP) and its association with apoptosis in congestive heart failure (CHF). BACKGROUND: Congestive heart failure is a heterogeneous syndrome with multiple hemodynamic, neuroendocrine and immune abnormalities. Although edematous CHF may be associated with endotoxemia and increased cytokine production, peripheral blood leukocyte functions in advanced CHF remain unclear. METHODS: Thirty patients with acute decompensated CHF (mean age [+/- SEM] 74.9 +/- 3.1 years) and 20 healthy controls underwent determination of MTP, intracellular oxidants and apoptosis in three subsets of peripheral blood leukocytes. The measurements were repeated after the time of recompensation. RESULTS: Patients with acute CHF showed marked MTP reduction and increased intracellular oxidant formation in three subsets of leukocytes upon entry into the study. These changes were more prominent in patients with peripheral edema. The decline in MTP was correlated with the severity of the peripheral edema and plasma concentration of cortisol, nitrogen metabolites and tumor necrosis factor-alpha (p < 0.01). After clinical stabilization, MTP gradually recovered. Leukocytes underwent increased propensity of apoptosis one week after the time of recompensation. CONCLUSIONS: The mitochondrial depolarization and apoptosis of leukocytes in decompensated heart failure suggest that CHF is associated with severity-dependent impairments in leukocyte function. Accentuated hormonal and cytokine abnormalities and increased circulating oxidants may contribute to these changes. Early and aggressive management of advanced heart failure is helpful in the recovery of these immune abnormalities.
Aged↗
Humic acid induced growth retardation in a sertoli cell line, TM4.
Humic acid (HA) is a fluorescent deep brown organic, polymeric compound composed of phenolic acid. Intraperitoneal injection of HA in rats induced testicular morphological changes including degeneration of the seminiferous tubule, reduction in the number of Sertoli cells and spermatogonia, and a loss of spermatids. It was suggested that Sertoli cells may be involved in the progression of testicular atrophy. In this study, we used a mouse Sertoli cell Line, TM4, to investigate the effect of HA on Sertoli cells and the mechanism of the testicular atrophy induced by HA. We found that the cell growth of TM4 cells were reduced in 1 to 4 days of HA exposure. FACScan analysis of the DNA content of HA-treated TM4 cells revealed that there was no sub-G1 peak, indicating that the TM4 cells did not commit to the programmed cell death. However, a large proportion of TM4 cells were arrested at the G1 phase. The percentage of TM4 cells at the G1 phase increased from 36% to 84% after HA treatment for 4 days. Western blot assay of HA-treated TM4 cells showed that the expression of cyclin D1 protein decreased while the expression of p27kiP1 protein increased. These results suggest that HA-induced testicular atrophy is linked in part to an inhibitory effect on the growth of Sertoli cells. This model may be useful in investigation of environmental agents inducing testicular atrophy.
Animals↗
Deletion mapping of chromosome 16q24 in hepatocellular carcinoma in Taiwan and mutational analysis of the 17-beta-HSD gene localized to the region.
Human chromosome band 16q24 commonly undergoes loss of heterozygosity (LOH) in human hepatocellular carcinoma (HCC). To further localize the region of deletion on 16q24 and to evaluate the genetic role of 17-beta-HSD, which is near 16q24, in HCC, we examined the pattern of loss of heterozygosity in 88 HCC patients. DNAs from 88 pairs of HCCs and corresponding non-tumor parts were prepared. Loss of heterozygosity on chromosomes 16q24 was investigated by 11 sets of microsatellite markers. Mutation analysis of type II 17-beta-HSD was performed by automatic sequencing. LOH on 16q24 for at least 1 locus was found in 43 of the 88 tumor DNAs (49%). Three non-overlapping regions of frequent LOH were defined in these 43 tumors with partial deletions. The first region was between D16S516 loci and D16S507, encompassed by a 1-cM region, defined by the D16S504. The second region was defined by the 17HSDB2 locus between D16S505 and D16S422, encompassed approximately by a 1-cM region. The third region was between D16S520 and D16S413, defined by D16S3048, encompassed approximately by a 4-cM region. Homozygous deletions of any exons in 17HSDB2 gene were identified in 7 of 27 cases (26%). Automated sequencing analysis of 17HSDB2 failed to demonstrate mutations in any of these specimens. Our data suggest that the 17HSDB2 locus is a frequent target of deletion in HCC but the inactivation of 17HSDB2 may not involve sequence mutations. Furthermore, the presence of the other 2 frequent LOH regions suggest that the putative tumor suppressor genes at these locations might be involved in the development of HCC.
17-Hydroxysteroid Dehydrogenases↗
Induction of oxidative stress by humic acid through increasing intracellular iron: a possible mechanism leading to atherothrombotic vascular disorder in blackfoot disease.
Humic acid (HA), a potential toxin that has penetrated the drinking well water of blackfoot disease-endemic areas in Taiwan, has been implicated as an etiological factor of this disease. In this study, we investigated the effects of HA on the generation of reactive oxygen species (ROS) in cultured human umbilical vein endothelial cells (HUVECs). The generation of ROS was monitored by flow cytometry. Pretreatment of HUVECs with HA induced reactive oxygen species in a dose- and time-dependent manner. Xanthine oxidase inhibitor (Allopurinol), NADPH oxidase inhibitor (diphenylene iodomium) and calcium chelator (BAPTA) could not reduce the generation of ROS. Protein kinase C inhibitor (H7) could reduce the generation of ROS slightly, but the intracellular antioxidant glutathione monoethyl ester and the iron chelator desferrioxamine (DFO) could inhibit the generation of ROS completely. HA also enhanced the expression of ferritin and induced intracellular chelatable iron; however, HA reduced the expression of transferrin receptor. Pretreatment with DFO inhibited HA-mediated increases of ferritin synthesis and intracellular chelatable iron, but caused recovery of the inhibitory effect on transferrin receptor. Cotreatment with iron and HA induced more ROS and intracellular chelatable iron than iron or HA treatment alone. Furthermore, HA enhanced the accumulation of iron in endothelial cells. These data demonstrate that HA can increase the generation of ROS through enhancing the accumulation of intracellular iron. Taken together, our findings suggest that iron mediates HA-associated oxidative stress in endothelial cells, which may be a possible mechanism leading to atherothrombotic vascular injury observed for patients with blackfoot disease.
Arterial Occlusive Diseases↗
Age-related changes in the mitochondrial depolarization induced by oxidative injury in human peripheral blood leukocytes.
Aging is associated with impaired immunity and reduced host defenses. Mitochondrial bioenergetic dysfunctions and reduced antioxidative ability of immunocompetent cells may contribute to this phenomenon. In this study, 60 healthy volunteers of different age groups donated their blood after overnight fasting. Leukocytes were subjected to oxidative injuries by exposure to t-butylhydroperoxide, and were labeled with fluorochromes for measuring mitochondria transmembrane potential (delta psi m), membrane peroxidation and mitochondrial oxidant formation. delta psi m declined after t-butylhydroperoxide exposure, and the change was more prominent in leukocytes from older individuals. Cyclosporin A partly restored delta psi m, implying the contributing role of mitochondrial permeability transition pores. The mitochondrial depolarization was accompanied by increased oxidant formation and oxidation of pyridine nucleotides, which were more prominent in older subjects. The results support the view that the bioenergetic functions of mitochondria are more susceptible to oxidative injury in aged individuals. The decreased ability of leukocytes to resist oxidative stress may contribute to immunosenescence in humans.
Adult↗
Enhanced oxidative stress and accelerated cellular senescence in glucose-6-phosphate dehydrogenase (G6PD)-deficient human fibroblasts.
Glucose-6-phosphate dehydrogenase (G6PD) is involved in the generation of reduced nicotinamide adenine dinucleotide phosphate (NADPH) and the maintenance of the cellular redox balance. The biological effects of G6PD deficiency in nucleated cells were studied using G6PD-deficient human foreskin fibroblasts (HFF). In contrast to that of normal HFF, the doubling time of G6PD-deficient cells increased readily from population doubling level (PDL) 15 to 63. This was accompanied by a significant increase in the percentage of G(1) cells. The slow-down in growth preceded an early entry of these cells into a nondividing state reminiscent of cellular senescence. These cells exhibited a significant increase in level of senescence-associated beta-galactosidase (SA-beta-gal) staining. The importance of G6PD activity in cell growth was corroborated by the finding that ectopic expression of active G6PD in the deficient cells prevented their growth retardation and early onset of senescence. Mechanistically, the enhanced fluorescence in dichlorofluorescin (H(2)DCF)-stained G6PD-deficient cells suggests the possible involvement of reactive oxygen species in senescence. Taken together, our results show that G6PD deficiency predisposes human fibroblasts to retarded growth and accelerated cellular senescence. Moreover, G6PD-deficient HFF provides a useful model system for delineating the effects of redox alterations on cellular processes.
Cell Cycle↗
Humic acid suppresses the LPS-induced expression of cell-surface adhesion proteins through the inhibition of NF-kappaB activation.
Humic acid (HA), a potential toxin when penetrating the drinking well water of blackfoot disease-endemic areas in Taiwan, has been implicated as one of the etiological factors of this disease. In this study, we investigated the effects of HA on the expression of human vascular endothelial-leukocyte adhesion molecules and the activation of nuclear factor kappa B (NF-kappaB) in cultured human umbilical vein endothelial cells (HUVECs). The expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin was monitored by flow cytometry. Pretreatment of HUVECs with HA inhibited the lipopolysaccharide (LPS)-induced expression of these three adhesion molecules in a dose- and time-dependent manner. Since NF-kappaB can regulate the expression of these adhesion molecules, NF-kappaB activation was assessed by electrophoretic mobility shift assay (EMSA). Our results reveal that the activation of NF-kappaB by LPS is suppressed by HA in a dose- and time-dependent manner. Furthermore, HA reduces NF-kappaB binding to DNA slightly, but completely inhibits the degradation of IkappaBalpha at a concentration of 100 microg/ml. Thus, all our data demonstrate that HA can inhibit the LPS-induced expression of adhesion molecules through the inhibition of NF-kappaB activation. HA may also suppress the immune or inflammatory reaction of HUVECs responsible for endotoxin, which could be one possible explanation for the causes of the infection and inflammation observed for patients with blackfoot disease. Our results also suggest that immune or inflammatory disturbance occurs for patients with blackfoot disease and that NF-kappaB may be a critical molecule in the pathogenesis of this disease.
Cell Adhesion↗
Cellular glucose-6-phosphate dehydrogenase (G6PD) status modulates the effects of nitric oxide (NO) on human foreskin fibroblasts.
Glucose-6-phosphate dehydrogenase (G6PD) plays an important role in cellular redox homeostasis, which is crucial for cell survival. In the present study, we found that G6PD status determines the response of cells exposed to nitric oxide (NO) donor. Treatment with NO donor, sodium nitroprusside (SNP), caused apoptosis in G6PD-deficient human foreskin fibroblasts (HFF1), whereas it was growth stimulatory in the normal counterpart (HFF3). Such effects were abolished by NO scavengers like hemoglobin. Ectopic expression of G6PD in HFF1 cells switched the cellular response to NO from apoptosis to growth stimulation. Experiments with 1H-¿1,2,4ŏxadiazolo¿4, 3-aquinoxalin-1-one and 8-bromo-cGMP showed that the effects of NO on HFF1 and HFF3 cells were independent of cGMP signalling pathway. Intriguingly, trolox prevented the SNP-induced apoptosis in HFF1 cells. These data demonstrate that G6PD plays a critical role in regulation of cell growth and survival.
Apoptosis↗
Humic acid-induced echinocyte transformation in human erythrocytes: characterization of morphological changes and determination of the mechanism underlying damage.
Blackfoot disease (BFD) is a peripheral arterial occlusive disease found among human inhabitants along the southwest coast of Taiwan. Well water used for drinking and cooking contains humic acid (HA), which may be a possible etiological factor. In this study, HA toxicity was investigated in human erythrocytes and was found to induce echinocytic formation. Morphological changes occurred in both a concentration- and time-dependent fashion. The presence of HA was also observed to facilitate the loading of erythrocytes with excess Ca(2+) (1 mM), which may have occurred following permeability changes in cell membranes, leading to echinocytic transformations. Sodium dodecyl sulfate (SDS) gel electrophoresis indicated that echinocyte formation was due to the oxidation of normal membrane proteins that were replaced by high-molecular-weight proteins. Humic acid also induced hemoglobin oxidation in erythrocytes. Data show that oxidative stress generated by HA as well as direct effects were exerted on the cytoskeleton of erythrocytes, and these may be significant factors in the etiology of BFD.
Anemia↗
The oxidative metabolism of circulating phagocytes in ankylosing spondylitis: determination by whole blood chemiluminescence.
OBJECTIVE: Superoxide anion radicals within the human body are regarded as a major cause of inflammation. However, their role in the pathogenesis of ankylosing spondylitis (AS) has not been well identified. This study aimed at investigating the relation between AS and the oxidative metabolism of phagocytes in whole blood. METHODS: 24 patients with classic AS were examined to determine their clinical status; complete blood count, erythrocyte sedimentation rate (ESR), and C reactive protein (CRP) were determined, and levels of the superoxide anion radicals in the patients with AS and 21 healthy subjects were assessed by the ultraweak chemiluminescence method. Subsequently, the relation between this disease and phagocytes was examined by using N-formyl-methionyl-leucyl-phenylalanine (fMLP) and phorbol-12-myristate-13-acetate (PMA) stimulants. RESULTS: In clinical assessments, patients with AS had abnormally raised serum CRP (>10 mg/l) and ESR (>15 mm/1st h) levels. In contrast with healthy subjects, patients with AS had significantly increased rates of superoxide anion radical production in their whole blood either in the resting state or with either fMLP or PMA stimulation. In addition, chemiluminescence maximum light intensity was significantly higher in patients with AS than in healthy subjects after fMLP or PMA stimulation. CONCLUSIONS: Our results suggest that the phagocytes of patients with AS are partly activated in the resting state, and are sensitive to fMLP or PMA stimulation. The priming of phagocytes in the bloodstream is likely to be a causative factor in the onset of AS.
Adult↗
Baicalein, a novel apoptotic agent for hepatoma cell lines: a potential medicine for hepatoma.
This study has demonstrated that baicalein has anticancer effectiveness against human hepatoma cells. The dose response of baicalein in Hep G2 and Hep J2 cells indicates that baicalein decreased viability >90%. In comparison, baicalein had only minimal effects on the viability of control Chang liver cells. Flow cytometric analysis showed that baicalein inhibited the cell cycle of Hep G2 cells in the S phase. In addition, baicalein treatment resulted in a decreased mitochondrial transmembrane potential and damaged the integrity of the cell membrane. The TdT-mediated dUTP-biotin nick end labeling assay results indicated that baicalein elicited a significant increase of DNA fragmentation in Hep G2 cells after incubation for 48 hours. These results indicate that baicalein is an effective antihepatoma agent with minimal influence on noncancer cells. The effects of baicalein on Hep G2 cells include inhibition of the S phase of the cell cycle, dysfunction of mitochondria, and initiation of apoptosis.
Apoptosis↗
Enchancing effect of patented whey protein isolate (Immunocal) on cytotoxicity of an anticancer drug.
To determine the enhancing effect of a whey protein isolate on the cytotoxicity of a potential anticancer drug, baicalein, the human hepatoma cell line Hep G2 was assigned to grow in different media for four days, and cell growth and apoptosis were investigated. The control group was grown in normal medium; the other three groups were grown in whey protein isolate (Immunocal) medium, baicalein medium, and a combination of Immunocal and baicalein. As indicated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, survival rate was significantly lower in cells grown in baicalein + Immunocal than in cells grown in baicalein alone. In contrast, there was no significant difference in survival rate of the cells grown in Immunocal. In the investigation of apoptosis, cells grown in baicalein + Immunocal showed a higher phosphatidylserine exposure, lower mitochondrial transmembrane potential, and nearly 13 times more cells undergoing apoptosis than cells grown in baicalein alone. We also demonstrated that Immunocal reduced glutathione (GSH) in Hep G2 cells by 20-40% and regulated the elevation of GSH, which was in response to baicalein. In conclusion, Immunocal seemed to enhance the cytotoxicity of baicalein by inducing more apoptosis; this increase in apoptotic cells may be associated with the depletion of GSH in Hep G2 cells. This is the first study to demonstrate, in vitro, that Immunocal may function as an adjuvant in cancer treatments.
Antineoplastic Agents↗
Methyl methanesulfonate and hydrogen peroxide differentially regulate p53 accumulation in hepatoblastoma cells.
Genotoxic chemicals not only damage cellular DNA, but may also induce cell apoptosis if they are lethal to the cell. p53, Bcl-2 and Bax play important roles in the regulation of genotoxic chemical induced cell apoptosis. Since the mechanisms by which cellular DNA damaged by different DNA-damaging chemicals may not be the same, we studied the involvement of p53, Bcl-2 and Bax in apoptosis induced by methyl methanesulfonate (MMS) and hydrogen peroxide (H2O2). H2O2 damages DNA by free radical generation and MMS damages DNA by DNA methylation. At non-lethal doses, both H2O2 and MMS induced high level of p53 protein accumulation. Nevertheless, while the amount of p53 protein increased with the dose of MMS and the occurrence of apoptotic cell death events, H2O2 doses that induce cell apoptosis attenuated the p53 protein accumulation level. Lethal MMS treatment also increased Bax, but not Bcl-2 expression, whereas in H2O2 induced apoptosis, the level of both Bcl-2 and Bax declined. These results indicate that toxic chemicals differentially regulate the accumulation of p53 protein. Thus, the pathways of toxic chemicals induced cell apoptosis are different and independent.
Apoptosis↗
Peroxisome proliferation, adipocyte determination and differentiation of C3H10T1/2 fibroblast cells induced by humic acid: induction of PPAR in diverse cells.
Humic acid, a high-molecular-weight polyphenolic compound, exists abundantly in soil, natural water, and various terrestrial and aquatic environments. Humic acid causes peroxisome proliferation in mouse liver and induces the expression of peroxisome proliferator activated receptor (PPAR) in BNL CL.2 cells. Both cytotoxicity and flow cytometry show that humic acid inhibits the growth of C3H10T1/2 cells at G1 phase. C3H10T1/2 fibroblast cells express PPARgamma and the adipocyte P2 (aP2) genes which convert into adipocytes after being treated with humic acid. Our findings may provide a unique model for studying the molecular control of determination and differentiation of mesodermal cell lineages.
Adipocytes↗
Simultaneous detection of vitamin C and uric acid by capillary electrophoresis in plasma of diabetes and in aqueous humor in acute anterior uveitis.
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Aqueous Humor↗
Oxidative injury induced by synthetic humic acid polymer and monomer in cultured rabbit articular chondrocytes.
Humic substance has been proposed as one of the causative factors of Kashin-Beck disease (KBD), an endemic osteoarthritic disorder with necrosis of chondrocytes widely prevalent in some regions of China. In order to exclude the complications of natural humic substance, here we prepared phenolic polymers of synthetic humic acid (SHA) by oxidation of phenolic monomer, the protocatechuic acid (PCA). The biological effects of SHA and PCA on primary culture of rabbit articular chondrocytes were investigated. We found that not only SHA but also PCA caused chondrocyte injury, as evidenced by the loss of cell viability measured with methylthiazol tetrazolium (MTT) assay and the increased release of intracellular lactate dehydrogenase (LDH). Both SHA and PCA could result in lipid peroxidation and glutathione (GSH) depletion in chondrocytes, indicating that oxidative stress may be involved in chondrocyte injury. Furthermore, a marked increase in intracellular calcium level ([Ca2+]i) occurred after chondrocytes treated with SHA or PCA. These results suggest that chondrocyte injury elicited by SHA or PCA may be mediated through the occurrence of oxidative stress and the disruption of intracellular Ca2+ homeostasis. Data also suggest that the monomeric phenolic acid may be considered one of the causative factors of KBD in addition to humic substance.
Animals↗