Phenethyl isothiocyanate promotes immune responses in normal BALB/c mice, inhibits murine leukemia WEHI-3 cells, and stimulates immunomodulations in vivo.

Enhanced cruciferous vegetable consumption is associated with the reduction of cancer incidence as shown in epidemiological studies. Phenethyl isothiocyanate (PEITC), one of the important compounds in cruciferous vegetables, has been shown to induce apoptosis in many types of human cancer cell lines, but there is no available information addressing the effects on normal and leukemia mice in vivo. The purpose of this study is to focus on the in vivo effects of PEITC on immune responses of normal and WEHI-3 leukemia BALB/c mice in vivo. Influences of PEITC on BALB/c mice after intraperitoneal (i.p.) injection with WEHI-3 cells and normal mice were investigated. In normal BALB/c mice, PEITC did not affect the body weight when compared to the olive oil treated animals. Moreover, PEITC promoted phagocytosis by macrophages from peripheral blood mononuclear cells (PBMC) and peritoneal cavity, increased the levels of CD11b and Mac-3, decreased the level of CD19 and promoted natural killer (NK) cell cytotoxic activity, but it did not alter the level of CD3. Also, PEITC enhanced T cell proliferation after concanavalin A (Con A) stimulation. Otherwise, PEITC increased the body weight, but decreased the weight of liver and spleen as compared to the olive oil-treated WEHI-3 leukemia mice. PEITC also increased the level of CD19, decreased the levels of CD3 and Mac-3 rather than influence in the level of CD11b, suggesting that the differentiation of the precursor of macrophages and T cells was inhibited, but the differentiation of the precursor of B cells was promoted in leukemia mice. Furthermore, PEITC enhanced phagocytosis by monocytes and macrophages from PBMC and peritoneal cavity, and also promoted the NK cell cytotoxic activity in comparison with the group of leukemia mice. Based on these observations, the biological properties of PEITC can promote immune responses in normal and WEHI-3 leukemia mice in vivo. © 2011 Wiley Periodicals, Inc. Environ Toxicol, 2013.

Diallyl disulfide induced signal transducer and activator of transcription 1 expression in human colon cancer colo 205 cells using differential display RT-PCR.

Diallyl disulfide is one of the components of garlic and has been demonstrated to induce apoptosis in many cancer cell lines, though it is not reported to be associated with signal transducer and activator of transcription 1 (STAT1) expression. Moreover the role of STAT1 does not directly affect apoptosis in cancer cells after exposure to chemotherapy agents, though some reports showed that STAT1 is associated with apoptosis. In this study, differential display RT-PCR was used to examine the effects of diallyl disulfide (DADS) on human colon cancer cells (colo 205). The results demonstrated that DADS induced the expression of STAT1 which was also confirmed using Western blotting. STAT1 decoy oligonucleotides were also used to block STAT1 mRNA and led to a decrease in the levels of STAT1 and to subsequence decrease in the percentage of apoptosis induced by DADS in examined colo 205 cells.

Curcumin-induced cell cycle arrest and apoptosis in human acute promyelocytic leukemia HL-60 cells via MMP changes and caspase-3 activation.

Curcumin (diferuloylmethane), is a natural product derived from the root of the plant Curcuma longa. For centuries, it has been used as a spice and as a herbal medicine in Chinese populations. Curcumin has been shown to inhibit cell proliferation, cell cycle arrest, cyclooxygenase (COX)-1 and -2 expression and apoptosis in several human cancer cell lines. The aim of this investigation was to clarify the mechanisms by which curcumin induced cytotoxicity and apoptosis in human leukemia HL-60 cells. The effects of curcumin on the levels of reactive oxygen species (ROS), Ca+2 production, cyclin E, cdc25c, wee1, Bcl-2, Bax, the changes of mitochondrial membrane potential (MMP), cytochrome c release and the activation of caspase-3 were also investigated in the HL-60 cells. Results of flow cytometry and DAPI staining assays indicated that curcumin induced cytotoxicity and apoptosis in the examined cells. The results from flow cytometry assay indicated that curcumin induced ROS and Ca+2 productions, decreased the levels of MMP and increased the activity of caspase-3, leading to cell apoptosis. Western blot assay also revealed that curcumin increased the levels of Bax and the release of cytochrome c, and decreased the levels of Bcl-2 in the examined cells. The inhibition of caspase-3 activation by z-VAD-fmk (broad-spectrum caspase inhibitor) completely blocked curcumin-induced apoptosis in HL-60 cells.

Involvement of Bax, Bcl-2, Ca2+ and caspase-3 in capsaicin-induced apoptosis of human leukemia HL-60 cells.

The role of Ca2+ on the effects of capsaicin on human leukemia HL-60 cells in vitro and the molecular mechanisms of capsaicin-induced apoptosis were investigated. The flow cytometric analysis indicated that capsaicin decreased the percentage of viable HL-60 cells, via the induction of G0/G1-phase cell cycle arrest and apoptosis. Capsaicin-induced G0/G1-phase arrest involved the suppression of CDK2 and the cyclin E complex, which are check-point enzymes for cells moving from G0/G1- to S-phase. Capsaicin-induced apoptosis was associated with the elevation of intracellular reactive oxygen species and Ca2+ production, decreased the levels of mitochondrial membrane potential, promoted cytochrome c release and increased the activation of caspase-3. An intracellular Ca2+ chelator (BAPTA) significantly inhibited capsaicin-induced apoptosis. Capsaicin-induced apoptosis was time-and dose-dependent. These results suggest that the capsaicin-induced apoptosis of HL-60 cells may result from the activation of caspase-3 and the intracellular Ca2+ release pathway.

Ellagic acid inhibits growth and arylamine N-acetyltransferase activity and gene expression in Staphylococcus aureus.

It is well documented that N-acetyltransferase (NAT) plays a key role in the N-acetylation of arylamine compounds. Ellagic acid was demonstrated to elicit dose-dependent bacteriostatic activity and inhibition of N-acetylation of 2-aminofluorene (AF). N-acetylation of AF in S. aureus was determined by high preformance liquid chromatography. The apparent values of Km and Vmax of NAT were decreased after co-treatment with 0.5 mM ellagic acid in the cytosol of S. aureus. PCR also indicated that ellagic acid inhibited NAT gene expression (NAT mRNA) in S. aureus.

Survey of CTX-M-3 extended-spectrum beta-lactamase (ESBL) among cefotaxime-resistant Serratia marcescens at a medical center in middle Taiwan.

Thirty-four clinical isolates of Serratia marcescens nonsusceptible to cefotaxime were collected from a medical center in middle Taiwan. Confirmatory tests for extended-spectrum beta-lactamases (ESBLs) by cefotaxime and ceftazidime +/- clavulanic acid using Etest ESBL Screen identified only one ESBL producer; the remaining 33 isolates revealed nondeterminable results, because of off-scale minimum inhibitory concentration (MIC) levels for cefotaxime +/- clavulanic acid. Agar microdilution method using broader MIC ranges confirmed 21 ESBL-producers and one non-determinable result, achieving a highly predicting value compared to golden standard by PCR and DNA sequencing analysis, which identified 22 (65%) isolates containing blaCTX-M-3 genes. Only one strain carried concurrent CTX-M-3 and SHV-5 conferring high-level MICs to both cefotaxime (128 microg/mL) and ceftazidime (64 microg/mL). Other enzymatic mechanisms, such as chromosome-encoded AmpC including a novel SRT-2 enzyme, may confer resistance to cefotaxime on the remaining 12 isolates without ESBL bla genes. Thus, it is unreliable to predict the resistance mechanism by antibiogram, and current Etest ESBL Screen tests. Our study highlights expanding efforts to detect ESBLs in S. marcescens are urgently needed in Taiwan.

Curcumin decreases the DNA adduct formation, arylamines N-acetyltransferase activity and gene expression in human colon tumor cells (colo 205).

The effects of curcumin on the N-acetyltransferase (NAT) activity, AF-DNA adduct formation and NAT gene expression were examined using the human colon tumor cell line (colo 205). Various concentrations of curcumin were added to the cytosols or to the medium of human colon tumor cells. The NAT activity was determined by high performance liquid chromatography assaying for the amounts of acetylated 2-aminofluorene (AAF) and p-aminobenzoic acid (N-Ac-PABA) and nonacetylated 2-aminofluorene (AF) and p-aminobenzoic acid (PABA). The NAT activity in the human colon tumor cells and cytosols was suppressed by curcumin in a dose-dependent manner. The results demonstrated that gene expression (NAT1 mRNA) in human colon tumor cells was inhibited by curcumin. The apparent values of Km and Vmax of NAT of human colon tumor cells were also decreased by curcumin in cytosols. Curcumin may act as a noncompetitive inhibitor. After the incubation of human colon tumor cells with AF with or without curcumin cotreatment, the cells were recovered and DNA was prepared, hydrolyzed to nucleotides, the adducted nucleotides were extracted into butanol and AF-DNA adducts analyzed by HPLC. The results also demonstrated that when curcumin was added to the media a decrease in AF-DNA adduct formation was seen in the human colon tumor cells. The finding of AF-DNA adduct formation in cultured human colon tumor cells suggests the usefulness of cultured cells for assessing arylamine-induced DNA damage.

Detection of cefotaxime-resistant CTX-M-3 in clinical isolates of Serratia marcescens.

Strains of Serratia marcescens (isolated in a hospital during April and August 2000) resistant to ampicillin, chloramphenicol, trimethoprim-sulfamethoxazole, streptomycin, tetracycline, and gentamicin were characterized. Out of a total of 34 clinical isolates 6 (17.6 %) exhibited the extended spectrum beta-lactamases (ESBL) resistance; they were also resistant to cefotaxime (minimum inhibitory concentration, MIC > or = 128 microg/mL) but susceptible to imipenem (MIC < or = 0.5 microg/mL). This multidrug resistance was shown to be transferred by a conjugative plasmid. Transconjugants revealed similar MIC profiles when compared to the parental strains. Isoelectric focusing revealed one major transferable beta-lactamase (pI 8.4) which was further identified as CTX-M-3 by PCR and gene sequencing. The presence of strains with this type of ESBL showed the evolution of bla genes and their dissemination among at least three species of the family Enterobacteriaceae isolated within a single hospital. The predominance of CTX-M type enzymes found in this area of Taiwan appeared to be similar to that described in Poland.

Benzyl isothiocyanate inhibits murine WEHI-3 leukemia cells in vitro and promotes phagocytosis in BALB/c mice in vivo.

Many evidences have shown that dietary intake of cruciferous vegetables could protect against the risk of various types of malignancies. Benzyl isothiocyanate (BITC), one of the compounds from cruciferous vegetables, had shown induced cell cycle arrest and apoptosis in cancer cells. However, there is no available information to address that BITC affects murine leukemia cells in vitro and in vivo. Here, we investigated in vitro effects of BITC on murine leukemia WEHI-3 cells. BITC decreased the percentage of viable cells via G0/G1 arrest and apoptosis in WEHI-3 cells. BITC induced apoptosis through the dysfunction of mitochondria (decreased the levels of mitochondria membrane potential) and activation of caspase-3. Then we investigated in vivo effects of BITC on murine leukemia WEHI-3 cells and the results indicated that BITC decreased the weights of liver and spleen and it also decreased the percentage of CD11b and Mac-3 markers, indicating that the differentiation of the precursor of macrophage and B cells was inhibited. BITC promoted the activity of macrophage phagocytosis in cells which are isolated from PBMC and peritoneal (i.p.). Taken together, BITC can affect WEHI-3 cells in vitro and in vivo.

Quercetin-induced apoptosis acts through mitochondrial- and caspase-3-dependent pathways in human breast cancer MDA-MB-231 cells.

There has been considerable evidence recently demonstrating the anti-tumour effects of flavonols. Quercetin, an ubiquitous bioactive flavonol, inhibits cells proliferation, induces cell cycle arrest and apoptosis in different cancer cell types. The precise molecular mechanism of quercetin-induced apoptosis in human breast cancer cells is unclear. The purpose of this study was to investigate effects of quercetin on cell viability and to determine its underlying mechanism in human breast cancer MDA-MB-231 cells. Quercetin decreased the percentage of viable cells in a dose- and time-dependent manner, which was associated with cell cycle arrest and apoptosis. Quercetin did not increase reactive oxygen species generation but increased cytosolic Ca(2+) levels and reduced the mitochondrial membrane potential (DeltaPsi(m)). Quercetin treatment promoted activation of caspase-3, -8 and -9 in MDA-MB-231 cells. Caspase inhibitors prevented the quercetin-induced loss of cell viability. Quercetin increased abundance of the pro-apoptotic protein Bax and decreased the levels of anti-apoptotic protein Bcl-2. Confocal laser microscope examination indicated that quercetin promoted apoptosis-inducing factor (AIF) release from mitochondria and stimulated translocation to the nucleus. Taken together, these findings suggest that quercetin results in human breast cancer MDA-MB-231 cell death through mitochondrial- and caspase-3-dependent pathways.

Berberine inhibits arylamine N-acetyltransferase activity and gene expression in Salmonella typhi.

The effects of berberine on growth, arylamine N-acetyltransferase (NAT) activity, and gene expression in Salmonella Typhi (Typhi) were described. The growth inhibition of Typhi was determined by measuring absorbance by optical density (OD at 650 nm). The NAT activity was determined by measuring the levels of 2-aminofluorene (AF) and N-acetyl-2-aminofluorene (AAF) by high-performance liquid chromatography. The results demonstrated that 24-h berberine treatment decreased bacteria growth and amounts of AAF in Typhi. Western blotting and flow cytometry were used for examining the levels of NAT after bacteria were cotreated with or without various concentrations of berberine, and results indicated that berberine decreased the levels of NAT in Typhi. Polymerase chain reaction was used for examining the gene expression of NAT (mRNA NAT), and results indicated that berberine affects mRNA NAT1 expression in Typhi.