Development and characterization of microsatellite markers from the yellow passion fruit (Passiflora edulis f. flavicarpa)

Here we described the development of the first set of Passiflora microsatellite loci isolated from an enriched genomic library. A sample of 43 individuals from 12 accessions of the yellow passion fruit was used to characterize those loci, which revealed up to 20 alleles per locus. Two loci were monomorphic. The observed (H_O) and expected (H_E) heterozygosities were very similar, as expected for a self‐incompatible species. Allelic diversity (H_T) was 0.444. This set of markers will permit genetic structure analyses of cultivated and wild populations of Passiflora, and contribute for integrating genetic maps based on dominant markers, as they can provide bridge alleles.     

Phenotypic Features of Circulating Leukocytes from Non-human Primates Naturally Infected with Trypanosoma cruzi Resemble the Major Immunological Findings Observed in Human Chagas Disease

Background

Cynomolgus macaques (Macaca fascicularis) represent a feasible model for research on Chagas disease since natural T. cruzi infection in these primates leads to clinical outcomes similar to those observed in humans. However, it is still unknown whether these clinical similarities are accompanied by equivalent immunological characteristics in the two species. We have performed a detailed immunophenotypic analysis of circulating leukocytes together with systems biology approaches from 15 cynomolgus macaques naturally infected with T. cruzi (CH) presenting the chronic phase of Chagas disease to identify biomarkers that might be useful for clinical investigations.

Methods and Findings

Our data established that CH displayed increased expression of CD32⁺ and CD56⁺ in monocytes and enhanced frequency of NK Granzyme A⁺ cells as compared to non-infected controls (NI). Moreover, higher expression of CD54 and HLA-DR by T-cells, especially within the CD8⁺ subset, was the hallmark of CH. A high level of expression of Granzyme A and Perforin underscored the enhanced cytotoxicity-linked pattern of CD8⁺ T-lymphocytes from CH. Increased frequency of B-cells with up-regulated expression of Fc-γRII was also observed in CH. Complex and imbricate biomarker networks demonstrated that CH showed a shift towards cross-talk among cells of the adaptive immune system. Systems biology analysis further established monocytes and NK-cell phenotypes and the T-cell activation status, along with the Granzyme A expression by CD8⁺ T-cells, as the most reliable biomarkers of potential use for clinical applications.

Conclusions

Altogether, these findings demonstrated that the similarities in phenotypic features of circulating leukocytes observed in cynomolgus macaques and humans infected with T. cruzi further supports the use of these monkeys in preclinical toxicology and pharmacology studies applied to development and testing of new drugs for Chagas disease.     

Helianthus tuberosus agglutinin directly induces neutrophil migration, which can be modulated/inhibited by resident mast cells.

We investigated the effect of Helianthus tuberosus agglutinin (HTA) on neutrophil migration in vivo and in vitro. The role of resident cells in this effect was analyzed. Peritonitis was induced by injecting stimuli into rat (150-200 g) peritoneal cavities, and in vitro neutrophil chemotaxis was performed using a Boyden microchamber. HTA (80, 200, or 500 microg/mL per cavity) induced significant in vivo neutrophil migration (p < 0.05); in vitro assays showed that this lectin also induced neutrophil chemotaxis, an effect inhibited by the incubation of lectin associated with alpha-D(+)-mannose, its specific binding sugar. Depletion of the resident-cell population by peritoneal lavage did not alter HTA-induced neutrophil migration (200 microg/mL per cavity). The opposite strategy, increasing peritoneal macrophages by intraperitoneally injecting rats with thioglycollate, did not enhance the neutrophil migration produced by HTA (200 microg/mL per cavity). In addition, injection of supernatant from HTA-stimulated macrophage culture (300 microg/mL) into rat peritoneal cavities did not induce neutrophil migration. However, reduction of the peritoneal mast-cell population potentiated the neutrophil migration (p < 0.05) induced by HTA (200 microg/mL per cavity). Lectin from H. tuberosus has a direct neutrophil chemotatic effect that is modulated by mast cells.     

Spitting out information: Trigona bees deposit saliva to signal resource locations

Stingless bees of the species Trigona spinipes (Fabricius 1793) use their saliva to lay scent trails communicating the location of profitable food sources. Extracts of the cephalic labial glands of the salivary system (not the mandibular glands, however) contain a large amount (approx. 74%) of octyl octanoate. This ester is also found on the scent-marked substrates at the feeding site. We demonstrate octyl octanoate to be a single compound pheromone which induces full trail following behaviour. The identification of the trail pheromone in this widely distributed bee makes it an ideal organism for studying the mechanism of trail following in a day flying insect.     

Antinociceptive and anti-inflammatory effects of electroacupuncture on experimental arthritis of the rat temporomandibular joint

This study investigated the antinociceptive and anti-inflammatory effects of electroacupuncture (EA) on zymosan-induced acute arthritis of the rat temporomandibular joint (TMJ). Male Wistar rats were injected with saline or zymosan (control group; 2?mg) into the left TMJ. Low frequency EA (10?Hz, 30?min) was performed at acupoints (LI4, LI11, ST36, ST44) or sham points 2?h after or 1?h before zymosan administration. Mechanical hypernociception was accessed by the electronic Von Frey method after zymosan administration. Rats were sacrificed 6?h after zymosan administration and the joint was removed for histopathological analysis, myeloperoxidase activity assessment, vascular permeability observations, and immunohistochemical verification of inflammatory mediators. The results showed that EA inhibited zymosan-induced hypernociception, compared with the control group and with the sham group (p?< 0.05). The results showed that EA inhibited inflammatory parameters such as neutrophil migration, vascular permeability, and tumour necrosis factor α (TNF-α), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) expression in the TMJ compared with the sham group (p < 0.05). Histopathological analysis showed that EA significantly inhibited edema and periarticular infiltration (p?< 0.05) compared with the control and sham groups. EA at acupoints produced antinociceptive and anti-inflammatory effects on zymosan-induced arthritis in the rat TMJ.     

Design,synthesis and in vitro evaluation against human cancer cells of 5-methyl-5-styryl-2,5-dihydrofuran-2-ones,a new series of goniothalamin analogues

The present work describes the preparation of a novel series of compounds based on the structure of goniothalamin (1), a natural styryl lactone with known cytotoxic and antiproliferative activities against a variety of cancer cell lines. A focused library of 17 goniothalamin analogues displaying the 5-methyl-2,5-dihydrofuran-2-one motif were prepared, and their cytotoxicity evaluated. While the analogues bearing methoxy and/or hydroxy groups on the aromatic moiety usually were at least three times less potent than the lead compound (1), ortho and para-trifluoromethyl analogues 10 and 11 exhibited levels of cytotoxicity similar to goniothalamin (1) against most cancer cell lines evaluated. One could suggest that the electronic effect of the trifluoromethyl group activates the inhibitor’s electrophilic site via reduction of the electron density of the α,β-unsaturated ester oxygen atom. These results provide new information on the structure activity relationship of these α,β-unsaturated styryl lactones, thereby further focusing the design of novel candidates.     

Efficacy of an injection of dinoprost tromethamine when given subcutaneously on luteal regression in lactating Holstein cows

The objectives of these studies were to evaluate the efficacy of a PGF(2alpha) (PGF) analog given through different routes on causing luteal regression in lactating dairy cows. In Experiment 1, lactating Holstein cows (n=118) at random stages of lactation were blocked by parity and days in milk (DIM) and, within each block, randomly assigned to receive PGF as an intra-muscular (IM) injection in the semimembranous/semitendinous muscle (CON), subcutaneous (SC) injection in the cervical area (SCN), or SC injection in the ischio-rectal fossa (IRF). Blood was sampled at 0, 12, 24, 36, and 48 h after treatment for assessment of progesterone concentration. In Experiment 2, a total of 379 lactating Holstein cows, 46+/-7 DIM, were blocked by DIM and, within each block, randomly assigned to receive treatment similar to CON or IRF groups from Experiment 1. Blood was sampled 0 and 48 h after treatment for assessment of progesterone concentration. Cows were classified as experiencing luteal regression when progesterone concentration was <1.0 ng/mL or <40% of initial concentration (0 h=100%). In Experiment 1, there was no effect of route of PGF treatment on decline in progesterone concentration and on the proportion of cows experiencing luteal regression by 12, 24, 36, and 48 h after treatment. Similarly, in Experiment 2, route of treatment did not affect either the decline in progesterone concentration or the proportion of cows that had luteal regression by 48 h after treatment. Treatment of lactating dairy cows with 25mg of PGF given SC in the ischio-rectal fossa did not affect either the decline in progesterone concentration or the proportion of cows that experienced luteal regression by 12, 24, 36, and 48 h after PGF treatment.     

Improved replication of the baculovirus Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) in vitro using proteins from Lonomia obliqua hemolymph

The baculovirus Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV), a member of the family Baculoviridae, has been widely applied as a biopesticide for the control of the velvetbean caterpillar, a pest of soybean crop field. Baculoviruses are considered safe and efficient agents for this purpose, because they do not infect vertebrates, being safe for the health of humans and animals, as well as to the environment. The objective of this work was to identify proteins obtained from Lonomia obliqua hemolymph with potential application in the optimization of baculovirus AgMNPV replication in Sf9 insect cell culture. In this work the improvement of the cell culture and viral replication of the AgMNPV baculovirus was observed when Grace medium was supplemented with 10 % (v/v) Fetal Bovine Serum (FBS), 1 % (v/v) hemolymph extract, or 3 % (v/v) of hemolymph fractions or hemolymph sub-fractions obtained by purifying hemolymph through High Performance Liquid Chromatography. Hemolymph presented a positive effect on the synthesis of polyhedra and enhanced baculovirus replication in Spodoptera frugiperda (Sf9) cells (TCID₅₀/mL), and led to Sf9 cell culture improvement. Grace medium supplemented with 10 % (v/v) FBS and 1 % (v/v) hemolymph provided an increase of baculovirus replication, when the cells were infected with multiplicity of infection of 1. In this case, the baculovirus replication was 6,443.91 times greater than that obtained with the control: Grace medium supplemented with 10 % (v/v) FBS. In addition, this work suggests that hemolymph from L. obliqua could have an interesting application in biotechnology, due to an increase in the viability of the cells and virus replication.     

Marked changes in endogenous antioxidant expression precede vitamin A-, C-, and E-protectable, radiation-induced reductions in small intestinal nutrient transport

Rapidly proliferating epithelial crypt cells of the small intestine are susceptible to radiation-induced oxidative stress, yet there is a dearth of data linking this stress to expression of antioxidant enzymes and to alterations in intestinal nutrient absorption. We previously showed that 5-14 days after acute γ-irradiation, intestinal sugar absorption decreased without change in antioxidant enzyme expression. In the present study, we measured antioxidant mRNA and protein expression in mouse intestines taken at early times postirradiation. Observed changes in antioxidant expression are characterized by a rapid decrease within 1h postirradiation, followed by dramatic upregulation within 4h and then downregulation a few days later. The cell type and location expressing the greatest changes in levels of the oxidative stress marker 4HNE and of antioxidant enzymes are, respectively, epithelial cells responsible for nutrient absorption and the crypt region comprising mainly undifferentiated cells. Consumption of a cocktail of antioxidant vitamins A, C, and E, before irradiation, prevents reductions in transport of intestinal sugars, amino acids, bile acids, and peptides. Ingestion of antioxidants may blunt radiation-induced decreases in nutrient transport, perhaps by reducing acute oxidative stress in crypt cells, thereby allowing the small intestine to retain its absorptive function when those cells migrate to the villus days after the insult.