Infection of mouse dermal fibroblasts by the monoxenous trypanosomatid protozoa Crithidia deanei and Herpetomonas roitmani

Traditionally, monoxenous trypanosomatid protozoa are not believed to infect vertebrate cells. Using light and electron microscopy, we show that the monoxenous trypanosomatids Crithidia deanei and Herpetomonas roitmani are able to infect dermal mouse fibroblasts in vitro. We present experimental evidence of phagocytosis of these trypanosomatids, and demonstrate their survival in vertebrate cells. This paper raises the question about the role of C. deanei and H. roitmani, and perhaps other monoxenous trypanosomatid species, in opportunistic infections of immunocompromised individuals and cutaneos lesions in vertebrate hosts.     

The factor H variant associated with age-related macular degeneration (His-384) and the non-disease-associated form bind differentially to C-reactive protein, fibromodulin, DNA, and necrotic cells

Recently, a polymorphism in the complement regulator factor H (FH) gene has been associated with age-related macular degeneration. When histidine instead of tyrosine is present at position 384 in the seventh complement control protein (CCP) domain of FH, the risk for age-related macular degeneration is increased. It was recently shown that these allotypic variants of FH, in the context of a recombinant construct corresponding to CCPs 6-8, recognize polyanionic structures differently, which may lead to altered regulation of the alternative pathway of complement. We show now that His-384, corresponding to the risk allele, binds C-reactive protein (CRP) poorly compared with the Tyr-384 form. We also found that C1q and phosphorylcholine do not compete with FH for binding to C-reactive protein. The interaction with extracellular matrix protein fibromodulin, which we now show to be mediated, at least in part, by CCP6-8 of FH, occurs via the polypeptide of fibromodulin and not through its glycosaminoglycan modifications. The Tyr-384 variant of FH bound fibromodulin better than the His-384 form. Furthermore, we find that CCP6-8 is able to interact with DNA and necrotic cells, but in contrast the His-384 allotype binds these ligands more strongly than the Tyr-384 variant. The variations in binding affinity of the two alleles indicate that complement activation and local inflammation in response to different targets will differ between His/His and Tyr/Tyr homozygotes.     

Analysis of recombinant human growth hormone by capillary electrophoresis with bilayer-coated capillaries using UV and MS detection

The characterization of recombinant human growth hormone (rhGH; somatropin) by capillary electrophoresis (CE) with UV-absorbance and mass spectrometric (MS) detection using capillaries noncovalently coated with polybrene (PB) and poly(vinyl sulfonic acid) (PVS) is demonstrated. Compared with bare fused-silica capillaries, PB-PVS coated capillaries yielded more favorable migration-time reproducibilities and higher separation efficiencies. Optimal separation conditions for the bilayer-coated capillaries comprised a background electrolyte (BGE) of 400 mM Tris phosphate (pH 8.5) yielding migration-time R.S.D.s of less than 1.0% and plate numbers above 300,000 for intact rhGH. The protein was also analyzed using the CE method described in the European Pharmacopoeia (Ph. Eur.) monograph. The pharmacopoeial method gave much longer analysis times (22 min versus 8 min), lower resolution and plate numbers, and consecutive shifts in migration time for rhGH, indicating possible interactions between the protein and the inner capillary wall. Due to stable migration times obtained with the coated capillaries, reliable profiling and quantification of rhGH and its byproducts in time was possible. Analysis of thermally degraded rhGH revealed the formation of two main degradation products. CE-mass spectrometry (MS) of this sample, using a PB-PVS coated capillary and a BGE of 75 mM ammonium formate (pH 8.5), suggests that these products are desamido forms of rhGH. Analyses of expired rhGH preparations with CE-UV and CE-MS indicated the presence of both deamidation and oxidation products.     

On measures of dissimilarity between point patterns: Classification based on prototypes and multidimensional scaling

This paper presents a collection of dissimilarity measures to describe and then classify spatial point patterns when multiple replicates of different types are available for analysis. In particular, we consider a range of distances including the spike‐time distance and its variants, as well as cluster‐based distances and dissimilarity measures based on classical statistical summaries of point patterns. We review and explore, in the form of a tutorial, their uses, and their pros and cons. These distances are then used to summarize and describe collections of repeated realizations of point patterns via prototypes and multidimensional scaling. We also show a simulation study to evaluate the performance of multidimensional scaling with two types of selected distances. Finally, a multivariate spatial point pattern of a natural plant community is analyzed through various of these measures of dissimilarity.     

Developmental aspects of the direct‐developing frog Adelophryne maranguapensis

Direct development in amphibians is characterized by the loss of aquatic breeding. The anuran Adelophryne maranguapensis is one example of a species with direct development, and it is endemic to the state of Ceará, Brazil. Detailed morphological features of A. maranguapensis embryos and the stages of sequential development have not been described before. Here, we analyzed all available genetic sequence tags in A. maranguapensis (tyr exon 1, pomc and rag1) and compared them with sequences from other species of Adelophryne frogs. We describe the A. maranguapensis reproductive tract and embryonic body development, with a focus on the limbs, tail, ciliated cells of the skin, and the egg tooth, which were analyzed using scanning electron microscopy. Histological analyses revealed ovaries containing oocytes surrounded by follicular cells, displaying large nuclei with nucleoli inside. Early in development, the body is unpigmented, and the neural tube forms dorsally to the yolk vesicle, typical of a direct‐developing frog embryo. The hindlimbs develop earlier than the forelimbs. Ciliated cells are abundant during the early stages of skin development and are less common during later stages. The egg tooth appears in the later stages and develops as a keratinized microridge structure. The developmental profile of A. maranguapensis presented here will contribute to our understanding of the direct‐development model and may help preserve this endangered native Brazilian frog. genesis 54:257–271, 2016. © 2016 Wiley Periodicals, Inc.     

Biotransformation of α-Bromoacetophenones by the Marine Fungus Aspergillus sydowii

The biotransformation reactions of α-bromoacetophenone (1), p-bromo-α-bromoacetophenone (2), and p-nitro-α-bromoacetophenone (3) by whole cells of the marine fungus Aspergillus sydowii Ce19 have been investigated. Fungal cells that had been grown in artificial sea water medium containing a high concentration of chloride ions (1.20 M) catalysed the biotransformation of 1 to 2-bromo-1-phenylethanol 4 (56%), together with the α-chlorohydrin 7 (9%), 1-phenylethan-1,2-diol 9 (26%), acetophenone 10 (4%) and phenylethanol 11 (5%) identified by GC-MS analysis. In addition, it was observed that the enzymatic reaction was accompanied by the spontaneous debromination of 1 to yield α-chloroacetophenone 5 (9%) and α-hydroxyacetophenone 6 (18%) identified by GC-FID analysis. When 2 and 3 were employed as substrates, various biotransformation products were detected but the formation of halohydrins was not observed. It is concluded that marine fungus A. sydowii Ce19 presents potential for the biotransformations of bromoacetophenone derivatives.     

Pristane, a non-antigenic adjuvant, induces MHC class II-restricted, arthritogenic T cells in the rat

Pristane-induced arthritis (PIA) in rats, a model for rheumatoid arthritis (RA), is a T cell-dependent disease. However, pristane itself is a lipid and unable to form a stable complex with a MHC class II molecule. Therefore, the specificity and function of the T cells in PIA are as unclear as in rheumatoid arthritis. In this study, we show that activated CD4+ alphabetaT cells, which target peripheral joints, transfer PIA. The pristane-primed T cells are of oligo or polyclonal origin as determined by their arthritogenicity after stimulation with several mitogenic anti-TCRVbeta and anti-TCRValpha mAbs. Arthritogenic cells secreted IFN-gamma and TNF-alpha (but not IL-4) when stimulated with Con A in vitro, and pretreatments of recipient rats with either anti-IFN-gamma or a recombinant TNF-alpha receptor before transfer ameliorated arthritis development. Most importantly, we show that these T cells are MHC class II restricted, because treatment with Abs against either DQ or DR molecules ameliorates arthritis development. The MHC class II restriction was confirmed by transferring donor T cells to irradiated recipients that were syngenic, semiallogenic, or allogenic to MHC class II molecules, in which only syngenic and semiallogenic recipients developed arthritis. These data suggest that the in vivo administration of a non-antigenic adjuvant, like pristane, activates CD4+ alphabetaT cells that are MHC class II restricted and arthritogenic.