Role of Intestinal Epithelial Cells in Immune Effects Mediated by Gram-Positive Probiotic Bacteria: Involvement of Toll-Like Receptors

The mechanisms by which probiotic bacteria exert their effects on the immune system are not completely understood, but the epithelium may be a crucial player in the orchestration of the effects induced. In a previous work, we observed that some orally administered strains of lactic acid bacteria (LAB) increased the number of immunoglobulin A (IgA)-producing cells in the small intestine without a concomitant increase in the CD4⁺ T-cell population, indicating that some LAB strains induce clonal expansion only of B cells triggered to produce IgA. The present work aimed to study the cytokines induced by the interaction of probiotic LAB with murine intestinal epithelial cells (IEC) in healthy animals. We focused our investigation mainly on the secretion of interleukin 6 (IL-6) necessary for the clonal expansion of B cells previously observed with probiotic bacteria. The role of Toll-like receptors (TLRs) in such interaction was also addressed. The cytokines released by primary cultures of IEC in animals fed with Lactobacillus casei CRL 431 or Lactobacillus helveticus R389 were determined. Cytokines were also determined in the supernatants of primary cultures of IEC of unfed animals challenged with different concentrations of viable or nonviable lactobacilli and Escherichia coli, previously blocked or not with anti-TLR2 and anti-TLR4. We concluded that the small intestine is the place where a major distinction would occur between probiotic LAB and pathogens. This distinction comprises the type of cytokines released and the magnitude of the response, cutting across the line that separates IL-6 necessary for B-cell differentiation, which was the case with probiotic lactobacilli, from inflammatory levels of IL-6 for pathogens.     

Shb null allele is inherited with a transmission ratio distortion and causes reduced viability in utero.

SHB is an Src homology 2 domain-containing adapter protein that has been found to be involved in numerous cellular responses. We have generated an Shb knockout mouse. No Shb-/- pups or embryos were obtained on the C57Bl6 background, indicating an early defect as a consequence of Shb- gene inactivation on this genetic background. Breeding heterozygotes for Shb gene inactivation (Shb+/-) on a mixed genetic background (FVB/C57Bl6/129Sv) reveals a distorted transmission ratio of the null allele with reduced numbers of Shb+/+ and Shb-/- animals, but increased number of Shb+/- animals. The Shb- allele is associated with various forms of malformations, explaining the relative reduction in the number of Shb-/- offspring. Shb-/- animals that were born were viable, fertile, and showed no obvious defects. However, Shb+/- female mice ovulated preferentially Shb- oocytes explaining the reduced frequency of Shb+/+ mice. Our study suggests a role of SHB during reproduction and development.     

Stimulation of acid formation by histamine, carbachol and pentagastrin in isolated pig parietal cells

Free cells were obtained by sequential incubations of pig gastric mucosa with pronase and collagenase. Approximately 10-15% of the cell population represented parietal cells. Accumulation of aminopyrine (AP) in the acid compartments of parietal cells was used as an index of their acid production. Histamine, carbachol and pentagastrin each independently stimulated aminopyrine accumulation. The initial rate of aminopyrine accumulation, observed after addition of 10(-4) M carbachol or 10(-6) M pentagastrin, were 32% and 10%, respectively, of that observed with 10(-4) M histamine. Steady-state aminopyrine accumulation in the presence of 10(-4) M histamine, 10(-4) M carbachol or 10(-6) M pentagastrin were 6.2 +/- 3.3, 2.6 +/- 0.6 and 3.0 +/- 1.5 pmol AP per 10(4) parietal cells, respectively (mean +/- SD, n = 5). The EC50 value for histamine was 3.4 +/- 1.4 X 10(-7) M, and for pentagastrin 5.9 +/- 4.2 X 10(-8) M (mean +/- SD, n = 5). The dose-response curve for carbachol was biphasic. A plateau was reached at 10(-5)-10(-4) M carbachol, and for this phase an apparent EC50 of 2.1 +/- 1.4 X 10(-6) M carbachol was calculated (mean +/- SD, n = 5). A further increase to 10(-3) M carbachol increased the aminopyrine accumulation. Atropine (10(-6) M) inhibited the response to concentrations up to 10(-4) M carbachol, but was without effect on the histamine- and pentagastrin-stimulation. The H2-receptor antagonist, cimetidine, right-shifted the dose--response curve for histamine. Also, the pentagastrin-stimulated aminopyrine accumulation was inhibited by cimetidine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regional occurrence of plasmid-mediated carbapenem-hydrolyzing oxacillinase OXA-58 in Acinetobacter spp. in Europe

The spread of the plasmid-mediated carbapenem-hydrolyzing oxacillinase OXA-58 was detected in Acinetobacter sp. clinical isolates from southern Europe, the Balkans, and central Turkey. It may contribute significantly to the emergence of carbapenem resistance in Acinetobacter spp., at least in this part of the world.     

Evaluation of Urea-motility-indole medium for recognition and differentiation of Salmonella and Shigella species in stool cultures.

A semisolid urea-motility-indole medium designed for detection in Enterobacteriaceae of urease activity, motility, and indole production in one tube was prepared and evaluated. The formulation of the medium was similar to that of Christensen urea agar, but the agar concentration was 0.2%, and 1% tryptone was added. Results with 687 strains of Enterobacteriaceae were the same as those obtained with standard test media (98% overall agreement). The urea-motility-indole medium was also used in combination with Kligler iron agar for the recognition and differentiation of Salmonella and Shigella species from colonies picked from plating media in fecal cultures. This combination was compared with the combination of Kligler iron agar and lysine iron agar with 507 strains of non-lactose-fermenting Enterobacteriaceae. Although both combinations enabled the presumptive recognition and differentiation of Salmonella and Shigella species, an analysis of data indicated that the combination of Kligler iron agar and urea-motility-indole medium performed better than the combination of Kligler iron agar and lysine iron agar in detecting Salmonella and Shigella species.     

Bacterial DNA activates human neutrophils by a CpG-independent pathway

Bacterial DNA stimulates macrophages, monocytes, B lymphocytes, NK cells, and dendritic cells in a CpG-dependent manner. In this work we demonstrate that bacterial DNA, but not mammalian DNA, induces human neutrophil activation as assessed by L-selectin shedding, CD11b upregulation, and stimulation of cellular shape change, IL-8 secretion, and cell migration. Induction of these responses is not dependent on the presence of unmethylated CpG motifs, as neutrophil stimulatory properties were neither modified by CpG-methylation of bacterial DNA nor reproduced by oligonucleotides bearing CpG motifs. We found that human neutrophils express Toll-like receptor (TLR) 9 mRNA. However, as expected for a CpG-independent mechanism, activation does not involve a TLR9-dependent signaling pathway; neutrophil stimulation was not prevented by immobilization of bacterial DNA or by wortmannin or chloroquine, two agents that inhibit TLR9 signaling. Of note, both single-stranded and double-stranded DNA were able to induce activation, suggesting that neutrophils might be activated by bacterial DNA at inflammatory foci even in the absence of conditions required to induce DNA denaturation. Our findings provide the first evidence that neutrophils might be alerted to the presence of invading bacteria through recognition of its DNA via a novel mechanism not involving CpG motifs.     

Immunohistochemical localization of laminin and type IV collagen in human cutaneous sensory nerve formations

We used immunohistochemical techniques and monoclonal antibodies to localize two basement membrane components (laminin and type IV collagen) in the nerves and sensory nerve formations, or corpuscles, supplying human digital skin. Furthermore, neurofilament proteins, S-100 protein and epithelial membrane antigen were studied in parallel. In dermal nerve trunks, immunostaining for laminin and type IV collagen was found to be co-localized in the perineurium and the Schwann cells, the stronger immunoreactivity being at the external surface of the cells. In the Meissner digital corpuscles, the immunoreactivity for laminin and type IV collagen was mainly observed underlying the cell surface of lamellar cells, while the cytoplasm was weakly immunolabelled or unlabelled. Finally, within Pacinian corpuscles co-localization of the two basement membrane molecules was encountered in the inner core, intermediate layer, outer core and capsule. Laminin and type IV collagen immunoreactivities were also found in blood vessels and sweat glands, apparently labelling basement membrane structures. The present results provide evidence for the presence of basement membrane in all periaxonic cells forming human cutaneous sensory nerve formations, and suggest that all of them are able to synthesize and release some basement membrane components, such as laminin and type IV collagen. The possible role of laminin in sensory nerve formations is discussed.     

Endophytic bacteria in Coffea arabica L

Eighty-seven culturable endophytic bacterial isolates in 19 genera were obtained from coffee plants collected in Colombia (n = 67), Hawaii (n = 17), and Mexico (n = 3). Both Gram positive and Gram negative bacteria were isolated, with a greater percentage (68%) being Gram negative. Tissues yielding bacterial endophytes included adult plant leaves, various parts of the berry (e.g., crown, pulp, peduncle and seed), and leaves, stems, and roots of seedlings. Some of the bacteria also occurred as epiphytes. The highest number of bacteria among the berry tissues sampled was isolated from the seed, and includes Bacillus , Burkholderia , Clavibacter , Curtobacterium , Escherichia , Micrococcus , Pantoea , Pseudomonas , Serratia , and Stenotrophomonas . This is the first survey of the endophytic bacteria diversity in various coffee tissues, and the first study reporting endophytic bacteria in coffee seeds. The possible role for these bacteria in the biology of the coffee plant remains unknown.     

Cesarean Delivery and Risk of Excess Weight Among Brazilian Preschool Children

Maternal and Child Health Journal - To study the relationship between cesarean Delivery (CD) and overweight in preschool children, considering the presence of birth aspects and demographic...     

Evaluation of the effect of a school garden as an educational didactic tool in vegetable and fruit consumption in teenagers

BACKGROUND/OBJECTIVESIncreasing the consumption of vegetables and fruits in Mexico remains a challenge. Promoting sustainable food production systems through schools may be an effective way to educate young people about food and nutrition issues. A study of nutritional education in adolescents, based on the school garden, is necessary in order to evaluate its effects on the consumption of fruits and vegetables among middle- and upper-income segments of the population. The objective of this study was to evaluate the effect of an educational intervention, accompanied by a school garden as an educational teaching tool, to improve vegetable and fruit consumption by Mexican teenagers attending a private middle/high school.SUBJECTS/METHODSTeenagers between 12 and 18 years of age (n = 126) attending a private middle/high school in Queretaro, Mexico participated in a 3-arm, controlled, comparative impact study using a vegetable and fruit consumption frequency questionnaire, food consumption diaries, a psychosocial factor assessment questionnaire of vegetable and fruit consumption, and structured interviews. The participants were randomized into 3 experimental groups: 1) food education + school garden (FE + SG), 2) FE only, and 3) control group (CG).RESULTSThe FE + SG and FE groups significantly increased the frequency and daily intake of vegetables and fruits compared to the CG. The FE + SG group showed greater understanding of, reflection upon, and analysis of the information they received about vegetable and fruit consumption, as well as a greater willingness to include these in their daily diet.CONCLUSIONSFE accompanied by a SG as a teaching tool is more effective at promoting vegetable and fruit consumption than either education alone or control in teenagers in middle-upper income segments of the population.