Eosinophil contamination of thioglycollate-elicited peritoneal macrophage cultures skews the functional readouts of in vitro assays

Thioglycollate-elicited peritoneal cells are a common source of macrophages for various in vitro assays, including stimulation with TLR ligands, cell signaling assays, phagocytosis, toxicology studies, and cytokine/chemokine production. The most common method for enrichment of cultured thioglycollate-elicited peritoneal cells is adherence. However, the presence of other cell types in freshly isolated and cultured thioglycollate-elicited peritoneal cells has not been examined. Here, we demonstrate that thioglycollate-elicited peritoneal cavity contains 55-60% nonmacrophage cells, and even after adherence, there are still 12-20% nonmacrophage cells remaining. Excluding macrophages, eosinophils are the major cell type in the freshly elicited cavity (30-40%). Eosinophils are also the major cell type contaminating in vitro cultures of thioglycollate-elicited peritoneal macrophages. Moreover, the contamination of macrophage cultures by eosinophils significantly diminishes activation of p38 MAPK and the serine threonine kinase Akt and production of proinflammatory cytokines in response to LPS stimulation. Taken together, these data suggest that thioglycollate-elicited peritoneal cells are far more heterogeneous than reported previously. Further, a failure to remove contaminating eosinophils may greatly affect the interpretation of results obtained with cultured thioglycollate-elicited macrophages. Thus, our data indicate that future studies intent on accurately assessing cultured macrophage phenotype and activation require depletion of all cocontaminating cells, especially eosinophils.     

Identification of human herpesviruses 1 to 8 in Tunisian multiple sclerosis patients and healthy blood donors

Members of the human Herpesviridae family are candidates for representing the macroenvironmental factors associated with multiple sclerosis (MS) pathogenesis. To verify the possible role of human herpesviruses (HHVs) as triggering or aggravating factors in relapsing–remitting multiple sclerosis clinical outcome, we studied the prevalence of all eight human herpesviruses in whole blood samples collected from 51 MS patients and from 51 healthy controls. The presence of DNA of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), varicella zoster virus (VZV), Epstein–Barr virus (EBV), human cytomegalovirus (HCMV), human herpesvirus 6 (HHV-6), human herpesvirus 7 (HHV-7) and human herpesvirus 8 (HHV-8) was searched by specific nested polymerase chain reaction. HHVs were significantly more prevalent in the blood of MS patients than in those of the controls (P < 10⁻⁴). HSV-1, HSV-2, HCMV and HHV-8 were negative in both MS patients and controls samples. In MS patients, EBV, HHV-7, HHV-6 and VZV were detected in 31.3%, 33.3%, 5.8% and 7.8% of samples, respectively, compared with 3.9%, 9.8%, 1.96% and 1.96%, respectively, of samples from controls. We found a statistically significant difference only for EBV DNA and for HHV-7 DNA prevalence (P < 0.001 and P = 0.03). Although these results indicate lack of apparent association in terms of gender, type of diagnosis, symptoms, disease score and β interferon treatment between EBV or HHV-7 to MS among Tunisian patients, heterogeneity related to genetic polymorphism as well as geographical distribution of the disease and of pathogens may be of significance.     

Neuroendocrine tumors of the lung: A five-year retrospective experience of Egyptian NCI (2010–2014)

Background

The spectrum of lung neuroendocrine tumors (NETs) encompasses low grade typical carcinoid (TC), intermediate grade atypical carcionid (AC) and high grade, both large cell neuroendocrine carcinoma (LCNEC) and small cell lung cancer (SCLC), with extreme differences in management and survival.

Fungal contamination, natural occurrence of mycotoxins and resistance for aflatoxin accumulation of some broad bean (Vicia faba L.) cultivars.

Using glucose-Czapek's and 10% NaCl glucose-Czapek's agar media incubated at 28 +/- 2 degrees C, nine genera and 18 species were identified from 10 different cultivars seed of broad bean. Aspergillus and Penicillium were the most common genera on the two types of media used. The most common fungal species were Aspergillus flavus, A. niger, A. tamarii and Penicillium chrysogenum. Thin-layer chromatographic analysis indicated that seeds of one cultivar only was naturally contaminated with aflatoxins B1, B2, G1 and G2. Nine mycotoxin-free cultivars of broad bean were examined for resistance or susceptibility for aflatoxin production when inoculated by Aspergillus flavus IMI 89717. Results obtained revealed that seeds of two cultivars were highly resistant, two cultivars showed partial resistance and the remaining were susceptible to the establishment of A. flavus and aflatoxin accumulation. The results did not show any relationship between morphological characters (colour, shape and size) and the amount of aflatoxin produced on the different broad bean seed cultivars. Also the results reveal absence of significant variation in the total nitrogen of the highly susceptible, partially resistant and highly resistant groups of broad bean seeds examined. Mean values of calcium, total phosphate and potassium of both seed coat and seed kernel of the susceptible seeds were low as compared to those of partially and highly resistant seed cultivars. The susceptible seeds contained higher levels of magnesium, zinc and sodium while the resistant seeds contained lower ones.     

Evaluation of nutritional value,characteristics, functional properties of <Emphasis Type="Italic">Cymodocea nodosa</Emphasis> and its benefits on health diseases

Background

Nutritional fact study has prime importance to make the species edible and commercially viable to the food consumers. This is the first report that indicates the chemical characterization, functional, antioxidant and antihypertensive properties of Cymodocea nodosa to evaluate its nutritional status.

Methods

Physico-chemical determination was determined by colorimetric and spectroscopic analysis. The functional and texture properties were evaluated since a desirable texture should be retained. Bioactive substances were determined by liquid chromatography-high resolution electrospray ionization mass spectrometry HPLC-DAD-ESI/MS2 analysis. Health benefit of this plant was highlighting by the antioxidant and antihypertensive potentials.

Results

Results showed that the seagrass powder was characterized by a high content of fibers (56.4%), the fatty acids profile was dominated by the oleic acid, which represents about 62.0% of the total fatty acids and the functional properties proved important values of swelling capacity (6.71?±?0.2) and water holding capacity (12.26?±?0.25), that were comparable to those of some foodstuffs. Finally, the physico-chemical analysis shows the wealth in phenolic compounds, that could be explained by the high antioxidant and antihypertensive ability which was concentration dependent.

Conclusion

The results from this study suggested that this marine plant could be utilized as a healthy food item for human consumption.

     

Nonclinical aspects of biopharmaceutical development: discussion of case studies at a PhRMA-FDA workshop

Robust assessments of the nonclinical safety profile of biopharmaceuticals are best developed on a scientifically justified, case-by-case basis, with consideration of the therapeutic molecule, molecular target, and differences/similarities between nonclinical species and humans (ICH S6). Significant experience has been gained in the 10 years ensuing since publication of the ICH S6 guidance. In a PhRMA-FDA-sponsored workshop, "Nonclinical Aspects of Biopharmaceutical Development," industry and US regulatory representatives engaged in exploration of current scientific and regulatory issues relating to the nonclinical development of biopharmaceuticals in order to share scientific learning and experience and to work towards establishing consistency in application of general principles and approaches. The proceedings and discussions of this workshop confirm general alignment of strategy and tactics in development of biopharmaceuticals with regard to such areas as species selection, selection of high doses in toxicology studies, selection of clinical doses, the conduct of developmental and reproductive toxicity (DART) studies, and assessment of carcinogenic potential. However, several important aspects, including, for example, appropriate use of homologues, nonhuman primates, and/or in vitro models in the assessment of risk for potential developmental and carcinogenic effects, were identified as requiring further scientific exploration and discussion.     

Comparison of toxicity and teratogenic effects of salen and vo-salen on chicken embryo

Objectives: The toxic and teratogenic effects of salen (C₁₆H₁₆N₂O₂) and salen vanadium oxide (VOS) (C₁₆H₁₄N₂O₃V) were evaluated against chicken embryos along with chicken hepatic and fibroblastic cells in vitro cultures. Methods: Salen and VOS complexes were injected in the following concentrations: 5, 10, 20, 40, 80, 160, and 300?μM/egg for salen and 7.5, 15, 75, 120, 150, 240, and 300?μM/egg for VOS. In order to screen for skeletal malformations, the alizarin red clearing and staining method was employed. For studying the cytotoxic effects of these compounds, hepatic and fibroblastic cells were cultured and treated. Results: Our results show that injecting salen with various concentrations leads to a significant increase in embryonic mortality. Skeletal and morphological malformations resulting from salen injections included ectopic viscera and club foot. Our results show that embryonic mortality increased relative to the control group. In addition, alizarin red staining showed skeletal malformations like deletion of caudal vertebrae. Discussion: Our comparison showed that salen was a stronger teratogen than VOS, which may be due roles of the vanadium element, whose derivatives show physiological particulars and at low concentrations plays anticancer specifications without toxic effect. Conclusion: Results show that chicken embryos were sensitive to the toxicity of salen and VOS, and these compounds can affect the growth and ossification of the chicken embryos. Moreover, the cytotoxicity of salen and VOS shows that the viability of both salen and VOS-treated cells significantly decreased in a dose-dependent manner.