Lactobacillus paracasei subsp. paracasei LC01 positively modulates intestinal microflora in healthy young adults

Lactobacillus paracasei subsp. paracasei LC01 (LC01) can tolerate intestinal stresses and has antioxidant activity. To evaluate the effect of the bacterium on human intestinal microflora, a randomized, double-blind, placebo-controlled human trial was carried out. Fifty-two healthy adult volunteers were randomized equally to two groups. One group consumed 12% (wt/vol) skimmed milk supplemented with 10¹⁰ CFU of LC01 each day for the 4-week treatment period, and then consumed placebo in the next treatment period, separated by a 2-week washout. The other group followed the reverse order. Group-specific real-time PCR and biochemical analyses was used to determine the intestinal bacterial composition of fecal samples collected at the end of every period, and the concentration of short-chain fatty acids and ammonia. A significant inhibition in fecal Escherichia coli and increase in Lactobacillus, Bifidobacterium, and Roseburia intestinalis were observed after consumption of LC01. Acetic acid and butyric acid were significantly higher in the probiotic stage and fecal ammonia was significantly lower. The results indicated a modulation effect of LC01 on the intestinal microflora of young adults, suggesting a beneficial effect on bowel health. LC01 may have potential value as a probiotic.     

A novel set of 223 EST-SSR markers in <Emphasis Type="Italic">Casuarina</Emphasis> L. ex Adans.: polymorphisms,cross-species transferability,and utility for commercial clone genotyping

Simple sequence repeat (SSR) markers are very useful for genetic applications in plants, but SSR resource for the important tree genus Casuarina L. ex Adans. is still limited. In this study, we report a novel set of 223 SSR markers in Casuarina developed from expressed sequence tag (EST) resource of GenBank. The 223 EST-SSR markers were polymorphic among 10 unrelated individuals of C. equisetifolia L. Johnson, with the number of alleles per locus (N_a), observed heterozygosity (H_o), expected heterozygosity (H_e), and polymorphic information content (PIC) averaging at 5.5, 0.72, 0.86, and 0.63, respectively. The rates of cross-species transferability ranged from 96.9% (C. glauca Sieber ex Sprengel) through 97.8% (C. cunninghamiana Miquel) to 99.1% (C. junghuhniana Miquel). Fifty-five C. equisetifolia clones widely planted in China were successfully genotyped with a subset of 20 EST-SSRs. These newly developed markers will have a great potential for genetic and breeding applications in Casuarina species and related taxa.     

Large-area gold nanohole arrays fabricated by one-step method for surface plasmon resonance biochemical sensing

Surface plasmon resonance (SPR) nanosensors based on metallic nanohole arrays have been widely reported to detect binding interactions in biological specimens. A simple and effective method for constructing nanoscale arrays is essential for the development of SPR nanosensors. In this work, we report a one-step method to fabricate nanohole arrays by thermal nanoimprinting in the matrix of IPS (Intermediate Polymer Stamp). No additional etching process or supporting substrate is required. The preparation process is simple, time-saving and compatible for roll-to-roll process, potentially allowing mass production. Moreover, the nanohole arrays were integrated into detection platform as SPR sensors to investigate different types of biological binding interactions. The results demonstrate that our one-step method can be used to efficiently fabricate large-area and uniform nanohole arrays for biochemical sensing.     

Associations Between Selenium Content in Hair and Kashin-Beck Disease/Keshan Disease in Children in Northwestern China: a Prospective Cohort Study

The purpose of this study is to explore more trace elements (zinc, potassium, copper, iron, boron, manganese, selenium, chromium and cadmium elements) in addition to calcium, magnesium, lead and arsenic in the sera of patients with schizophrenia and the general population in China and to determine the effect of selenium on schizophrenia patients. Participants were collected from the Pingyin County Mental Health Hospital and Pingyin County. A t test was used to analyse the differences between schizophrenia patients and healthy subjects, and element content differences in gender. Logistic multivariate regression analysis was applied to analyse the influence of elements to schizophrenia. Repeated measures analysis of variance was used to analyse differences in the elements after 1 and 3 months. Mn, Se, Cd, Pb, Ca, Cu and Fe were lower than those in the normal group, and B, Cr, As, K and Mg were higher than those in the control group. The odd ratios (ORs) of serum As and serum K were 2.624 and 1.035, respectively. The ORs of sera Cr, Mn, Se, Ca and Cu were all below one. After intervention of ‘selenium weikang’ about 1 and 3 months, the serum As was decreased and the serum selenium and copper were increased. Cr, Mn, Se, Ca and Cu might have beneficial, statistically significant effects on schizophrenia. Elements As and K might be harmful to schizophrenia with statistical significance. After selenium supplementation, the schizophrenia patients improved in some factors, like the appetite and memory, and the As element decreased.     

Mutagenesis of benzo[a]pyrene diol epoxide in yeast: requirement for DNA polymerase zeta and involvement of DNA polymerase eta

Benzo[a]pyrene is a potent environmental carcinogen, which can be metabolized in cells to the DNA damaging agent anti-benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (anti-BPDE). We hypothesize that mutations induced by BPDE DNA adducts are mainly generated through an error-prone translesion synthesis that requires a specialized DNA polymerase (Pol). Using an in vivo mutagenesis assay in the yeast model system, we have examined the potential roles of Pol(zeta) and Pol(eta) in (+/-)-anti-BPDE-induced mutagenesis. In cells proficient in mutagenesis, (+/-)-anti-BPDE induced 85% base substitutions with predominant G --> C followed by G --> T transversions, 9% deletions of 1-3 nucleotides, and 6% insertions of 1-3 nucleotides. In rad30 mutant cells lacking Pol(eta), (+/-)-anti-BPDE-induced mutagenesis was reduced and accompanied by a moderate decrease in base substitutions and more significant decrease in deletions and insertions of 1-3 nucleotides. In rev3 mutant cells lacking Pol(zeta), (+/-)-anti-BPDE-induced mutagenesis was mostly abolished, leading to a great decrease in both base substitutions and deletions/insertions of 1-3 nucleotides. In contrast, large deletions/insertions were significantly increased in cells lacking Pol(zeta). Consistent with the in vivo results, purified yeast Pol(zeta) performed limited translesion synthesis opposite (+)- and (-)-trans-anti-BPDE-N(2)-dG DNA adducts with predominant G incorporation opposite the lesion. These results show that (+/-)-anti-BPDE-induced mutagenesis in yeast requires Pol(zeta) and partially involves Pol(eta) and suggest that Pol(zeta) directly participates in nucleotide insertions opposite the lesion, while Pol(eta) significantly contributes to deletions and insertions of 1-3 nucleotides.     

Peptide models of four possible insulin folding intermediates with two disulfides

The single-chain insulin (PIP) can spontaneously fold into native structure through preferred kinetic intermediates. During refolding, pairing of the first disulfide A20-B19 is highly specific, whereas pairing of the second disulfide is likely random because two two-disulfide intermediates have been trapped. To get more details of pairing property of the second disulfide, four model peptides of possible folding intermediates with two disulfides were prepared by protein engineering, and their properties were analyzed. The four model peptides were named [A20-B19, A7-B7]PIP, [A20-B19, A6-B7]PIP, [A20-B19, A6-A11]PIP, and [A20-B19, A7-A11]PIP according to their remaining disulfides. The four model peptides all adopt partially folded structure with moderate conformational differences. In redox buffer, the disulfides of the model peptides are more easily reduced than those of the wild-type PIP. During in vitro refolding, the reduced model peptides share similar relative folding rates but different folding yields: The refolding efficiency of the reduced [A20-B19, A7-A11]PIP is about threefold lower than that of the other three peptides. The present results indicate that the folding intermediates corresponding to the present model peptides all adopt partially folded conformation, and can be formed during PIP refolding, but the chance of forming the intermediate with disulfide [A20-B19, A7-A11] is much lower than that of forming the other three intermediates.     

Type I collagen-induced MMP-2 activation coincides with up-regulation of membrane type 1-matrix metalloproteinase and TIMP-2 in cardiac fibroblasts

Migration of cardiac fibroblasts is implicated in infarct healing and ventricular remodeling. Activation of matrix metalloproteinases induced by three-dimensional type I collagen, the principal component of the myocardial interstitium, is hypothesized to be essential for this migration. By utilizing primary cultures of cardiac fibroblasts and collagen lattice models, we demonstrated that type I collagen induced MMP-2 activation, and cells undergoing a change from isometric tension to mechanical unloading were associated with increased levels of total and active MMP-2 species. The collagen-induced MMP-2 activation coincided with up-regulated cellular levels of both membrane type 1-matrix metalloproteinase (MT1-MMP) and TIMP-2. A fraction of cellular membrane prepared from cells embedded in the collagen lattice containing active MT1-MMP and TIMP-2 was capable of activating pro-MMP-2, and exogenous TIMP-2 had a biphasic effect on this membrane-mediated MMP-2 activation. Interestingly, the presence of 43-kDa MT1-MMP species in a fraction of intracellular soluble proteins prepared from monolayer cells but not cells embedded in the lattices indicates that MT1-MMP metabolizes differently under the two different culture conditions. Treatment of cells embedded in the lattice with furin inhibitor attenuated pro-MT1-MMP processing and MMP-2 activation and impeded cell migration and invasion. These results suggest that the migration and invasion of cardiac fibroblasts is furin-dependent and that the active species of MT1-MMP and MMP-2 may be involved in both events.     

MicroRNA-200a Regulates Grb2 and Suppresses Differentiation of Mouse Embryonic Stem Cells into Endoderm and Mesoderm

The mechanisms by which microRNAs (miRNAs) affect cell fate decisions remain poorly understood. Herein, we report that miR-200a can suppress the differentiation of mouse embryonic stem (ES) cells into endoderm and mesoderm. Interestingly, miR-200a directly targets growth factor receptor-bound protein 2 (Grb2), which is a key adaptor in the Erk signaling pathway. Furthermore, high levels of miR-200a dramatically decrease Grb2 levels and suppress the appearance of mesoderm and endoderm lineages in embryoid body formation, as well as suppressing the activation of Erk. Finally, Grb2 supplementation significantly rescues the miR-200a-induced layer-formation bias and the Erk suppression. Collectively, our results demonstrate that miR-200a plays critical roles in ES cell lineage commitment by directly regulating Grb2 expression and Erk signaling.