Astragaloside IV enhances taxol chemosensitivity of breast cancer via caveolin-1-targeting oxidant damage

Accumulating evidence suggests that caveolin-1 (CAV-1) is a stress-related oncotarget and closely correlated to chemoresistance. Targeting CAV-1 might be a promising strategy to improve chemosensitivity for breast cancer treatment. Astragaloside IV (AS-IV), a bioactive compound purified from Astragalus membranaceus, has been shown to exhibit multiple bioactivities, including anticancer. However, the involved molecular targets are still ambiguous. In this study, we investigated the critical role of CAV-1 in mediating the chemosensitizing effects of AS-IV to Taxol on breast cancer. We found that AS-IV could enhance the chemosensitivity of Taxol with minimal direct cytotoxicity on breast cancer cell lines MCF-7 and MDA-MB-231, as well as the nontumor mammary epithelial cell line MCF-10A. AS-IV was further demonstrated to aggravate Taxol-induced apoptosis and G2/M checkpoint arrest. The phosphorylation of mitogen-activated protein kinase (MAPK) signaling extracellular signal-regulated kinase (ERK) and c-Jun N-terminal Kinase (JNK), except p38, was also abrogated by a synergistic interaction between AS-IV and Taxol. Moreover, AS-IV inhibited CAV-1 expression in a dose-dependent manner and reversed CAV-1 upregulation induced by Taxol administration. Mechanism study further demonstrated that AS-IV treatment triggered the eNOS/NO/ONOO⁻ pathway via inhibiting CAV-1, which led to intense oxidant damage. CAV-1 overexpression abolished the chemosensitizing effects of AS-IV to Taxol by inhibiting oxidative stress. In vivo experiments further validated that AS-IV increased Taxol chemosensitivity on breast cancer via inhibiting CAV-1 expression, followed by activation of the eNOS/NO/ONOO⁻ pathway. Taken together, our findings not only suggested the potential of AS-IV as a promising candidate to enhance chemosensitivity, but also highlighted the significance of CAV-1 as the target to reverse cancer drug resistance.     

MicroRNA-125b Confers the Resistance of Breast Cancer Cells to Paclitaxel through Suppression of Pro-apoptotic Bcl-2 Antagonist Killer 1 (Bak1) Expression

Paclitaxel (Taxol) is an effective chemotherapeutic agent for treatment of cancer patients. Despite impressive initial clinical responses, the majority of patients eventually develop some degree of resistance to Taxol-based therapy. The mechanisms underlying cancer cells resistance to Taxol are not fully understood. MicroRNA (miRNA) has emerged to play important roles in tumorigenesis and drug resistance. However, the interaction between the development of Taxol resistance and miRNA has not been previously explored. In this study we utilized a miRNA array to compare the differentially expressed miRNAs in Taxol-resistant and their Taxol-sensitive parental cells. We verified that miR-125b, miR-221, miR-222, and miR-923 were up-regulated in Taxol-resistant cancer cells by real-time PCR. We further investigated the role and mechanisms of miR-125b in Taxol resistance. We found that miR-125b was up-regulated in Taxol-resistant cells, causing a marked inhibition of Taxol-induced cytotoxicity and apoptosis and a subsequent increase in the resistance to Taxol in cancer cells. Moreover, we demonstrated that the pro-apoptotic Bcl-2 antagonist killer 1 (Bak1) is a direct target of miR-125b. Down-regulation of Bak1 suppressed Taxol-induced apoptosis and led to an increased resistance to Taxol. Restoring Bak1 expression by either miR-125b inhibitor or re-expression of Bak1 in miR-125b-overexpressing cells recovered Taxol sensitivity, overcoming miR-125-mediated Taxol resistance. Taken together, our data strongly support a central role for miR-125b in conferring Taxol resistance through the suppression of Bak1 expression. This finding has important implications in the development of targeted therapeutics for overcoming Taxol resistance in a number of different tumor histologies.     

Effect of enzyme and ligand protonation on the binding of folates to recombinant human dihydrofolate reductase: implications for the evolution of eukaryotic enzyme efficiency.

There is marked pH dependence of the rate constant (koff) for tetrahydrofolate (H4folate) dissociation from its ternary complex with human dihydrofolate reductase (hDHFR) and NADPH. Similar pH dependence of H4folate dissociation from the ternary complex of a variant of hDHFR with the substitution Phe31----Leu (F31L hDHFR) causes this dissociation to become rate limiting in the enzyme mechanism at pH approximately 5, and this accounts for the marked decrease in kcat for this variant as the pH is decreased from 7 to 5. This decreased kcat at low pH is not seen for most DHFRs. koff for dissociation of folate, dihydrofolate (H2folate), and H4folate from their binary complexes with hDHFR is similarly pH dependent. For all the complexes examined, the pH dependence of koff in the range pH 5-7 is well described by a pKa of about 6.2 and must be due to ionization of a group on the enzyme. In the higher pH range (7-10), koff increases further as the pH is raised, and this relation is governed by a second pKa which is close to the pKa for ionization of the amide group (HN3-C4O) of the respective ligands. Thus, ionization of the ligand amide group also increases koff. Evidence is presented that the dependence of pH on koff for hDHFR accounts for the shape of the kcat versus pH curve for both hDHFR as well as its F31L variant and contributes to the higher efficiency of hDHFR compared with bacterial DHFR.     

Selective N-alkylation of β-alanine facilitates the synthesis of a poly(amino acid)-based theranostic nanoagent

The development of functional amino acid-based polymeric materials is emerging as a platform to create biodegradable and nontoxic nanomaterials for medical and biotechnology applications. In particular, facile synthetic routes for these polymers and their corresponding polymeric nanomaterials would have a positive impact in the development of novel biomaterials and nanoparticles. However, progress has been hampered by the need to use complex protection-deprotection methods and toxic phase transfer catalysts. In this study, we report a facile, single-step approach for the synthesis of an N-alkylated amino acid as an AB-type functional monomer to generate a novel pseudo-poly(amino acid), without using the laborious multistep, protection-deprotection methods. This synthetic strategy is reproducible, easy to scale up, and does not produce toxic byproducts. In addition, the synthesized amino acid-based polymer is different from conventional linear polymers as the butyl pendants enhance its solubility in common organic solvents and facilitate the creation of hydrophobic nanocavities for the effective encapsulation of hydrophobic cargos upon nanoparticle formation. Within the nanoparticles, we have encapsulated a hydrophobic DiI dye and a therapeutic drug, Taxol. In addition, we have conjugated folic acid as a folate receptor-targeting ligand for the targeted delivery of the nanoparticles to cancer cells expressing the folate receptor. Cell cytotoxicity studies confirm the low toxicity of the polymeric nanoparticles, and drug-release experiments with the Taxol-encapsulated nanoparticles only exhibit cytotoxicity upon internalization into cancer cells expressing the folate receptor. Taken together, these results suggested that our synthetic strategy can be useful for the one-step synthesis of amino acid-based small molecules, biopolymers, and theranostic polymeric nanoagents for the targeted detection and treatment of cancer.     

High Bak Expression Is Associated with a Favorable Prognosis in Breast Cancer and Sensitizes Breast Cancer Cells to Paclitaxel

Breast cancer has become the leading cause of cancer-related death among women. A large number of patients become resistant to drug chemotherapy. Paclitaxel (Taxol) is an effective chemotherapeutic agent used to treat cancer patients. Taxol has been widely used in human malignancies including breast cancer because it can stabilize microtubules resulting in cell death by causing an arrest during the G2/M phase of the cell cycle. Pro-apoptotic Bcl-2 antagonist killer 1 (Bak) plays an important role in Taxol-induced apoptosis in breast cancer. In our present study, we investigated the expression of the Bak protein and clinicopathological correlations in a large sample of breast cancer tissues by immunohistochemistry. We found that the percentage of high scores of Bak expression in breast cancer was significantly lower than that of the non-cancerous breast control tissue. In addition, lower Bak expression was positively associated with the clinical TNM stage of breast cancer with a significant decrease in overall survival compared with those with higher Bak expression especially in the Luminal and HER2 subtypes. Importantly, higher Bak expression predicted a favorable clinical outcome in the cases treated with Taxol indicated by a higher overall survival than that of patients with lower Bak expression especially in Luminal and HER2 subtypes. Furthermore, these results were confirmed in vitro since overexpression of Bak sensitized breast cancer cells to Taxol by inhibiting proliferation and promoting apoptosis; in contrast, downregulation of Bak through siRNA transfection inhibited Taxol induced-apoptosis. Therefore, our results demonstrate that Bak acts as a sensitive biomarker and favorable prognostic factor for Taxol treatment in breast cancer. The restoration of Bak expression would be therapeutically beneficial for Taxol resistant breast cancer patients.     

Folate-conjugated iron oxide nanoparticles for solid tumor targeting as potential specific magnetic hyperthermia mediators: synthesis, physicochemical characterization, and in vitro experiments

New folate-conjugated superparamagnetic maghemite nanoparticles have been synthesized for the intracellular hyperthermia treatment of solid tumors. These ultradispersed nanosystems have been characterized for their physicochemical properties and tumor cell targeting ability, facilitated by surface modification with folic acid. Preliminary experiments of nanoparticles heating under the influence of an alternating magnetic field at 108 kHz have been also performed. The nanoparticle size, surface charge, and colloidal stability have been assessed in various conditions of ionic strength and pH. The ability of these folate "decorated" maghemite nanoparticles to recognize the folate receptor has been investigated both by surface plasmon resonance and in folate receptor expressing cell lines, using radiolabeled folic acid in competitive binding experiments. The specificity of nanoparticle cellular uptake has been further investigated by transmission electron microscopy after incubation of these nanoparticles in the presence of three cell lines with differing folate receptor expression levels. Qualitative and quantitative determinations of both folate nanoparticles and nontargeted control nanoparticles demonstrated a specific cell internalization of the folate superparamagnetic nanoparticles.     

SYNTHESIS OF FOLATE RECEPTOR-TARGETED AND DOXORUBICIN-COUPLED CHEMOTHERAPEUTIC NANOCONJUGATE AND RESEARCH INTO ITS MEDICAL APPLICATIONS

Folate-targeted drug delivery has become an alternative therapy for the treatment of various cancers. Folate receptors are known to be responsible for cellular accumulation of folate and folate analogs with high binding affinity. The anthracycline antibiotic doxorubicin has a broad spectrum of antineoplastic action and a correspondingly widespread degree of clinical use. In this work, we aimed to prepare a folate receptor-targeted doxorubicin delivery system to achieve minimal effect of doxorubicin on healthy cells and more cytotoxicity of it on tumor cells. Folate–poly(ethylene glycol)–doxorubicin (FOL-PEG-DOX) nanoconjugate was synthesized through this aim and characterized with nuclear magnetic resonance (NMR), zetasizer, and atomic force microscopy (AFM). Doxorubicin release studies were also performed in vitro. The size of FOL-PEG-DOX was 78.84 nm. The results indicated that doxorubicin release rate from the conjugate was faster at pH 5.0 than pH 7.4 and the amide bond between DOX and PEG was more stable at pH 7.4 than pH 5.0. As a consequence, FOL-PEG-DOX nanoconjugate could be a potentially useful delivery system for folate receptor-positive cancer cells.     

Kinetic evidence for two interconvertible forms of the folate transport protein from Lactobacillus casei.

Lactobacillus casei cells contain a folate transport protein which exhibits a high affinity for folate. The dissociation constant (KD) for folate derived from binding parameters at the steady state (at 0 degree C) is 0.4 nM at pH 7.5 and 0.1 nM at pH 6.0. In the present study, folate binding to this protein at pH 7.5 (and 0 degree C) was shown to follow second-order kinetics and to proceed with an association constant (k+1) of 4.9 X 10(7) liter/mol per min. K+1 was not affected by preincubation conditions which alter the energetic state of the cell. Measurements on the extent of binding showed further that (at 0 degree C) essentially all unoccupied folate-binding sites reside at or are readily accessible to the outer surface of the membrane. In contrast, after saturating the binding site with [3H]folate, the first-order rate constant (k-1) for dissociation of the bound substrate (at 0 degree C) was found to vary substantially with the conditions employed. k-1 was 0.028/min in freshly harvested cells, but it increased by 2.8-fold in cells preincubated at 23 degrees C for 60 min and by 5.4-fold in isolated membranes. In addition, the faster rate observed in preincubated cells (k-1 = 0.077/min) returned to a slower rate after brief exposure of the cells to pH 6.0 (k-1 = 0.041/min), glucose (k-1 = 0.050/min), or both (k-1 = 0.012/min). k-1 was twofold lower at pH 6.0 than at pH 7.5 and was less dependent on the preincubation conditions, although it also increased substantially (5.5-fold) when the cells were converted to plasma membranes. The proposed explanation for these results is that folate transport protein of L. casei exists in two forms which can be distinguished by the accessibility of the binding site to the external medium and whose amounts are dependent upon the presence of bound folate, the pH, and the energetic state of the cell. It is suggested that these forms are transport proteins with binding sites oriented towards the inner and outer surfaces of the membrane.     

p53 acetylation enhances Taxol-induced apoptosis in human cancer cells

Microtubule inhibitors (MTIs) such as Taxol have been used for treating various malignant tumors. Although MTIs have been known to induce cell death through mitotic arrest, other mechanisms can operate in MTI-induced cell death. Especially, the role of p53 in this process has been controversial for a long time. Here we investigated the function of p53 in Taxol-induced apoptosis using p53 wild type and p53 null cancer cell lines. p53 was upregulated upon Taxol treatment in p53 wild type cells and deletion of p53 diminished Taxol-induced apoptosis. p53 target proteins including MDM2, p21, BAX, and β-isoform of PUMA were also upregulated by Taxol in p53 wild type cells. Conversely, when the wild type p53 was re-introduced into two different p53 null cancer cell lines, Taxol-induced apoptosis was enhanced. Among post-translational modifications that affect p53 stability and function, p53 acetylation, rather than phosphorylation, increased significantly in Taxol-treated cells. When acetylation was enhanced by anti-Sirt1 siRNA or an HDAC inhibitor, Taxol-induced apoptosis was enhanced, which was not observed in p53 null cells. When an acetylation-defective mutant of p53 was re-introduced to p53 null cells, apoptosis was partially reduced compared to the re-introduction of the wild type p53. Thus, p53 plays a pro-apoptotic role in Taxol-induced apoptosis and acetylation of p53 contributes to this pro-apoptotic function in response to Taxol in several human cancer cell lines, suggesting that enhancing acetylation of p53 could have potential implication for increasing the sensitivity of cancer cells to Taxol.     

Functional characterization of PCFT/HCP1 as the molecular entity of the carrier-mediated intestinal folate transport system in the rat model

Proton-coupled folate transporter/heme carrier protein 1 (PCFT/HCP1) has recently been identified as a transporter that mediates the translocation of folates across the cellular membrane by a proton-coupled mechanism and suggested to be the possible molecular entity of the carrier-mediated intestinal folate transport system. To further clarify its role in intestinal folate transport, we examined the functional characteristics of rat PCFT/HCP1 (rPCFT/HCP1) expressed in Xenopus laevis oocytes and compared with those of the carrier-mediated folate transport system in the rat small intestine evaluated by using the everted tissue sacs. rPCFT/HCP1 was demonstrated to transport folate and methotrexate more efficiently at lower acidic pH and, as evaluated at pH 5.5, with smaller Michaelis constant (K(m)) for the former (2.4 microM) than for the latter (5.7 microM), indicating its characteristic as a proton-coupled folate transporter that favors folate than methotrexate as substrate. rPCFT/HCP1-mediated folate transport was found to be inhibited by several but limited anionic compounds, such as sulfobromophthalein and sulfasalazine. All these characteristics of rPCFT/HCP1 were in agreement with those of carrier-mediated intestinal folate transport system, of which the K(m) values were 1.2 and 5.8 microM for folate and methotrexate, respectively, in the rat small intestine. Furthermore, the distribution profile of the folate transport system activity along the intestinal tract was in agreement with that of rPCFT/HCP1 mRNA. This study is the first to clone rPCFT/HCP1, and we successfully provided several lines of evidence that indicate its role as the molecular entity of the intestinal folate transport system.     

紫杉醇抑制前列腺癌增殖的体内实验研究

目的探讨紫杉醇对人前列腺癌细胞PC-3增殖的体内抑制作用。方法建立体内绿色荧光蛋白（GFP）标记的人雄激素非依赖性前列腺癌细胞PC-3裸鼠原位移植瘤模型,观察紫杉醇对裸鼠前列腺癌原位移植瘤的体积、重量的影响。结果裸鼠模型体内实验显示,与对照组（100μL生理盐水）相比,紫杉醇处理组（0.5 mg/kg）在给药第18天后能显著抑制前列腺肿瘤的体积（P〈0.05）;紫杉醇处理组在抑制前列腺肿瘤重量方面与对照组相比亦有明显抑制作用（P〈0.05）。与对照组相比G31P处理组VEGF（P〈0.05）的表达差异具有统计学意义（免疫组化法）。结论紫杉醇在体内实验中能明显抑制人雄激素非依赖性前列腺癌细胞系PC-3的增殖。     

Effects of cultivation conditions on folate production by lactic acid bacteria

A variety of lactic acid bacteria were screened for their ability to produce folate intracellularly and/or extracellularly. Lactococcus lactis, Streptococcus thermophilus, and Leuconostoc spp. all produced folate, while most Lactobacillus spp., with the exception of Lactobacillus plantarum, were not able to produce folate. Folate production was further investigated in L. lactis as a model organism for metabolic engineering and in S. thermophilus for direct translation to (dairy) applications. For both these two lactic acid bacteria, an inverse relationship was observed between growth rate and folate production. When cultures were grown at inhibitory concentrations of antibiotics or salt or when the bacteria were subjected to low growth rates in chemostat cultures, folate levels in the cultures were increased relative to cell mass and (lactic) acid production. S. thermophilus excreted more folate than L. lactis, presumably as a result of differences in the number of glutamyl residues of the folate produced. In S. thermophilus 5,10-methenyl and 5-formyl tetrahydrofolate were detected as the major folate derivatives, both containing three glutamyl residues, while in L. lactis 5,10-methenyl and 10-formyl tetrahydrofolate were found, both with either four, five, or six glutamyl residues. Excretion of folate was stimulated at lower pH in S. thermophilus, but pH had no effect on folate excretion by L. lactis. Finally, several environmental parameters that influence folate production in these lactic acid bacteria were observed; high external pH increased folate production and the addition of p-aminobenzoic acid stimulated folate production, while high tyrosine concentrations led to decreased folate biosynthesis.     

Dismutation of dihydrofolate by dihydrofolate reductase

Degradation of 7,8-dihydrofolate (H2folate) in the presence of dihydrofolate reductase (DHFR) has been shown due not to an oxygenase activity of the reductase as previously reported but to dismutation of H2folate to folate and 5,6,7,8-tetrahydrofolate (H4folate). The reaction can be followed spectrophotometrically or by analysis of the reaction mixture by high-performance liquid chromatography (HPLC). The products have also been isolated and characterized. Oxygen uptake during the reaction is much less than stoichiometric with H2folate disappearance and is attributed to autoxidation of the H4folate formed. The dismutation activity is a property of highly purified Streptococcus faecium DHFR isoenzyme 2 (but not isoenzyme 1) and of Lactobacillus casei DHFR, but not of bovine liver DHFR. The activity is dependent on tightly bound NADP+ and/or NADPH. Removal of the nucleotide results in loss of dismutation activity, which is restored by adding NADP+ or NADPH. Maximum activity is obtained when approximately 1 mol equiv of nucleotide is added per mol of DHFR. It is proposed that in the dismutation reaction bound NADP(H) is alternately reduced and oxidized by incoming molecules of H2folate with release of folate and H4folate, respectively. The relatively slow rate of folate formation presumably limits the rate of the overall reaction. The equilibrium constant for the dismutation reaction is 19.4 +/- 7.4 at 22 degrees C and pH 7.0. Calculation of standard oxidation-reduction potentials at pH 7 gave values of -0.230 V for the H2folate/H4 folate pair and -0.268 V for the folate/H2folate pair. The mechanism by which NADP+ is retained by the enzyme from some sources during purification procedures is unclear.(ABSTRACT TRUNCATED AT 250 WORDS)     

Binding of radiolabeled folate and 5-methyltetrahydrofolate to cow's milk folate binding protein at pH 7.4 and 5.0. Relationship to concentration and polymerization equilibrium of the purified protein

Binding of folate (pteroylglutamate) and 5-methyltetrahydrofolate, the major endogenous form of folate, to folate binding protein purified from cow's milk was studied at 7°C to avoid degradation of 5-methyltetrahydrofolate. Both folates dissociate rapidly from the protein at pH 3.5, but extremely slowly at pH 7.4, most likely due to drastic changes in protein conformation occurring after folate binding. Dissociation of 5-methyltetrahydrofolate showed no increase at 37°C suggesting that protein-bound-5-methyltetrahydrofolate is protected against degradation. Binding displayed two characteristics, positive cooperativity and a binding affinity that increased with decreasing concentrations of the protein. The binding affinity of folate was somewhat greater than that of 5-methyl tetrahydrofolate, in particular at pH 5.0. Ligand-bound protein exhibited concentration-dependent polymerization (8-mers formed at 13 M) at pH 7.4. At pH 5.0, only folate-bound forms showed noticeable polymerization. The fact that folate at pH 5.0 surpasses 5-methyltetrahydrofolate both with regard to binding affinity and ability to induce polymerization suggests that ligand binding is associated with conformational changes of the protein which favor polymerization.     

Biological evaluation of new antitumor taxoids: Alteration of substitution at the C-7 and C-10 of docetaxel

A series of new docetaxol analogues have been designed and synthesized. And their cytotoxicities against cancer cells have been evaluated by MTT method. Most of these compounds showed selective inhibitions on human cancer cell lines. Among them, compound 8 exhibited higher inhibitory activity than Paclitaxel (Taxol) against several cancer cell lines. This work indicated that appropriate modification at C-7 and C-10 of docetaxel might be a promising approach for this unique class of anticancer compounds.     

Crystal structures of recombinant human dihydrofolate reductase complexed with folate and 5-deazafolate

The 2.3-A crystal structure of recombinant human dihydrofolate reductase (EC 1.5.1.3, DHFR) has been solved as a binary complex with folate (a poor substrate at neutral pH) and also as a binary complex with an inhibitor, 5-deazafolate. The inhibitor appears to be protonated at N8 on binding, whereas folate is not. Rotation of the peptide plane joining I7 and V8 from its position in the folate complex permits hydrogen bonding of 5-deazafolate's protonated N8 to the backbone carbonyl of I7, thus contributing to the enzyme's greater affinity for 5-deazafolate than for folate. In this respect it is likely that bound 5-deazafolate furnishes a model for 7,8-dihydrofolate binding and, in addition, resembles the transition state for folate reduction. A hypothetical transition-state model for folate reduction, generated by superposition of the DHFR binary complexes human.5-deazafolate and chicken liver.NADPH, reveals a 1-A overlap of the binding sites for folate's pteridine ring and the dihydronicotinamide ring of NADPH. It is proposed that this binding-site overlap accelerates the reduction of both folate and 7,8-dihydrofolate by simultaneously binding substrate and cofactor with a sub van der Waals separation that is optimal for hydride transfer.     

Rho GTPases RhoA and Rac1 Mediate Effects of Dietary Folate on Metastatic Potential of A549 Cancer Cells through the Control of Cofilin Phosphorylation

Folate, an important nutrient in the human diet, has been implicated in cancer, but its role in metastasis is not established. We have shown previously that the withdrawal of medium folate leads to the inhibition of migration and invasion of A549 lung carcinoma cells. Here we have demonstrated that medium folate regulates the function of Rho GTPases by enabling their carboxyl methylation and translocation to plasma membrane. Conversely, the lack of folate leads to the retention of these proteins in endoplasmic reticulum. Folate also promoted the switch from inactive (GDP-bound) to active (GTP-bound) GTPases, resulting in the activation of downstream kinases p21-activated kinase and LIM kinase and phosphorylation of the actin-depolymerizing factor cofilin. We have further demonstrated that in A549 cells two GTPases, RhoA and Rac1, but not Cdc42, are immediate sensors of folate status: the siRNA silencing of RhoA or Rac1 blocked effects of folate on cofilin phosphorylation and cellular migration and invasion. The finding that folate modulates metastatic potential of cancer cells was confirmed in an animal model of lung cancer using tail vein injection of A549 cells in SCID mice. A folate-rich diet enhanced lung colonization and distant metastasis to lymph nodes and decreased overall survival (35 versus 63 days for mice on a folate-restricted diet). High folate also promoted epithelial-mesenchymal transition in cancer cells and experimental mouse tumors. Our study provides experimental evidence for a mechanism of metastasis promotion by dietary folate and highlights the interaction between nutrients and metastasis-related signaling.     

Folate production by bifidobacteria as a potential probiotic property

The ability of 76 Bifidobacterium strains to produce folate was investigated. In order to evaluate folic acid productivity, bifidobacteria were cultivated in the folate-free semisynthetic medium SM7. Most of the tested strains needed folate for growth. The production and the extent of vitamin accumulation were not a function of species but were distinctive features of individual strains. Six strains among the 17 that grew without folate produced significantly higher concentrations of vitamin (between 41 and 82 ng ml(-1)). The effects of exogenous folate and p-aminobenzoic acid (PABA) concentrations on folate production were evaluated. In contrast to most of the other strains, the folate yield of B. adolescentis MB 239 was not negatively affected by either PABA or exogenous folic acid. Folate production by B. adolescentis MB 239 was studied in the pH range of the colonic environment, and a comparison of folate production on raffinose, lactose, and fructo-oligosaccharides, which belong to three important groups of fermentable intestinal carbon sources, was established. Differences in folate biosynthesis by B. adolescentis MB 239 were not observed as a function either of the pH or of the carbon source. Fecal culture experiments demonstrated that the addition of B. adolescentis MB 239 may increase the folate concentration in the colonic environment.     

Esomeprazole overcomes paclitaxel-resistance and enhances anticancer effects of paclitaxel by inducing autophagy in A549/Taxol cells

Non-small-cell lung cancer (NSCLC) is one of the most common malignancies, and the occurrence of drug-resistance severely limits the efficacy of anticancer drugs in the treatment of NSCLC. Identification of new agents to reverse drug-resistance in NSCLC treatment is of great importance and urgency both clinically and scientifically. In the present study, we found that A549/Taxol cells displayed a high level of resistance to paclitaxel with the resistance index up to 231. Importantly, esomeprazole could potentiate the antiproliferative effect of paclitaxel in A549/Taxol cells, but not in A549 cells. Further exploration on the underlying mechanisms revealed that esomeprazole decreased the intracellular pH via inhibiting V-ATPase expression in A549/Taxol cells. Meanwhile, esomeprazole pretreatment significantly promoted paclitaxel-induced polymerization of tubulin and enhanced the proportion of G2/M-arrested cells in A549/Taxol cells. Unfortunately, esomeprazole could only result in a slight decrease in the expression of P-gp in A549/Taxol cells. Interestingly, esomeprazole significantly increased paclitaxel-induced apoptosis, which was impeded by the autophagy inhibitor 3-MA in A549/Taxol cells. Taken together, our data suggest that esomeprazole is a promising chemosensitizer against paclitaxel-resistant NSCLC by inducing autophagy. Our study also offers a new strategy to solve the paclitaxel-resistance problem during NSCLC treatment.     

Micronutrient intake and risk of colon and rectal cancer in a Danish cohort

Background: Micronutrients may protect against colorectal cancer. Especially folate has been considered potentially preventive. However, studies on folate and colorectal cancer have found contradicting results; dietary folate seems preventive, whereas folic acid in supplements and fortification may increase the risk. Objective: To evaluate the association between intake of vitamins C, E, folate and beta-carotene and colorectal cancer risk, focusing on possibly different effects of dietary, supplemental and total intake, and on potential effect modification by lifestyle factors. Design: In a prospective cohort study of 56,332 participants aged 50–64 years, information on diet, supplements and lifestyle was collected through questionnaires. 465 Colon and 283 rectal cancer cases were identified during follow-up. Incidence rate ratios of colon and rectal cancers related to micronutrient intake were calculated using Cox proportional hazard analyses. Results: The present study found a protective effect of dietary but not supplemental folate on colon cancer. No association with any other micronutrient was found. Rectal cancer did not seem associated with any micronutrient. For both colon and rectal cancer, we found an interaction between dietary folate and alcohol intake, with a significant, preventive effect among those consuming above 10 g alcohol/day only. Conclusions: This study adds further weight to the evidence that dietary folate protects against colon cancer, and specifies that there is a source-specific effect, with no preventive effect of supplemental folic acid. Further studies should thus take source into account. Vitamins C, E and beta-carotene showed no relation with colorectal cancer.