Identification of Tumor Antigens in the HLA Peptidome of Patient-derived Xenograft Tumors in Mouse

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Highlights

• •Sufficient tumor tissues are often unavailable large HLA peptidome discovery.
• •Using patient derived xenograft (PDX) tumors can overcome this limitation.
• •The large PDX HLA peptidomes expand significantly those of the original biopsies.
• •The HLA peptidomes of the PDX tumors included many tumor antigens.

     

Study of some parameters which affect xylanase production: Strain selection, enzyme extraction optimization, and influence of drying conditions

Xylanases are glycosidases mainly responsible for the hydrolysis of β-1,4 linkages in xylan. Xylanase was produced in this work by solid-state fermentation using agro industrial residues with Aspergillus niger strain, which was screened through qualitative and quantitative methods. Extraction processes with different solvents were evaluated. Solvent volume, time, and agitation speed (shaker) were optimized using statistical designs. Drying studies of the solid fermented material were also conducted in a laboratory oven where the following conditions were applied: 42°C and 50°C for 20 h and 80°C for 1 h; 50°C and 75°C for 6 and 3 h, respectively. Best extraction conditions were 37 mL of solvent composed by NaCl solution (0.9%) with Tween 80 (0.1%) in 3 g of cultured material with agitation at 133 rpm in shaker for 4 min. Highest xylanase activity was 2,327 IU/gdm. The drying at 42°C for 20 h provided a better maintenance of xylanase activity (58% of xylanase activity).     

Lead accumulation in the aquatic fern Azolla filiculoides.

In this study, we characterized lead (Pb2+) accumulation and storage by the aquatic fern Azolla filiculoides. Lead precipitates were detected in the vacuoles of mesophyll cells of Azolla plants cultured for 6 d in rich growth medium containing 20 mg l(-1) Pb2+. Energy dispersive spectroscopy (EDS) analysis of the relative element content of leaves collected from these plants revealed a 100% increase in the levels of P, S, Na and Ca and a 40% decrease in Mg and Cl compared to the untreated plants. Both Azolla whole plants and isolated apoplasts were incubated for 6 d in 20 mg l(-1) Pb2+. Lead content in the whole plant composed 0.37%, 2.3% and 1.8% of the dry weight after 2, 4 and 6 d of growth, respectively, while the isolated Azolla apoplast contained 0.125%, 1.22% and 1.4% Pb2+, respectively. Lead content in Azolla whole plant increase by 200%, 100% and 22% after 2, 4 and 6 d of growth, respectively, when compared to Azolla apoplast. Dark, electron dense deposits of lead were observed in light and transmission electron microscope in leaf cells treated with lead. All the observed lead deposits were localized in vacuoles while larger lead deposits were found in mature leaves than in young leaves. No lead deposits were found in cells of the cyanobiont Anabaena when the plants were exposed to similar conditions. Activity and content of V-H+-ATPase were studied in Azolla plants grown in the presence of 20, 40 and 80 mg l(-1) of lead for a period of 4 d. Activity of V-H+-ATPase was increased by 190%, 210% and 220%, respectively, but the content of V-H+-ATPase was reduced by all lead concentrations.     

Automated identification of RNA conformational motifs: theory and application to the HM LSU 23S rRNA

We develop novel methods for recognizing and cataloging conformational states of RNA, and for discovering statistical rules governing those states. We focus on the conformation of the large ribosomal subunit from Haloarcula marismortui. The two approaches described here involve torsion matching and binning. Torsion matching is a pattern-recognition code which finds structural repetitions. Binning is a classification technique based on distributional models of the data. In comparing the results of the two methods we have tested the hypothesis that the conformation of a very large complex RNA molecule can be described accurately by a limited number of discrete conformational states. We identify and eliminate extraneous and redundant information without losing accuracy. We conclude, as expected, that four of the torsion angles contain the overwhelming bulk of the structural information. That information is not significantly compromised by binning the continuous torsional information into a limited number of discrete values. The correspondence between torsion matching and binning is 99% (per residue). Binning, however, does have several advantages. In particular, we demonstrate that the conformation of a large complex RNA molecule can be represented by a small alphabet. In addition, the binning method lends itself to a natural graphical representation using trees.     

Stand microclimate and physiological activity of tree leaves in an oak-hornbeam forest

Summary In an uneven-aged, multi-species oak-hornbeam forest at Báb, SW Slovakia (former IBP Forest Research Site), a series of micrometeorological and ecophysiological measurements started in 1985. The aims of the work are to improve understanding of physiological processes (photosynthesis, respiration, and transpiration) of adult trees and stand microclimate, to collect data for simulation of the canopy (stand) photosynthesis and for ecological synthesis of the functioning of the forest ecosystem. In this paper, photosynthetically active radiation (PAR), air temperature (AT) and relative humidity (RH), wind speed (WS), and CO₂ concentration ([CO₂]) in and above the forest are characterized for the fully leaved season, using diurnal courses, vertical profiles and isodiagrams (isopleths). Approximately 50% of incident PAR was absorbed by the upper 4–5 m layer of leaves and only approximately 5% or less penetrated to the forest floor. Vertical gradients of AT and RH were generally low, but large differences in diurnal ranges of AT and RH were observed between vertical levels. The upper leaf canopy greatly reduced WS, and at a height of about 14 m above the ground it was close to zero. The highest diurnal [CO₂] maximum and variations occurred at 1 m above the ground, and the lowest above the forest. In good light conditions in the forest, the entire leaf canopy (overstorey and understorey canopy) is a large sink of CO₂. At night the forest stand is a source of CO₂, the largest internal source being the soil and forest floor.     

Factors associated with consent for organ donation: a retrospective population-based study

Background:Optimizing the approach to and consent of potential organ donors maximizes patient autonomy and the availability of organs for transplants. We set out to identify modifiable factors associated with donation consent.Methods:We conducted a retrospective cohort study of consecutive adults (≥ 18 yr) referred for organ donation in Ontario between April 2013 and June 2019. We analyzed patient clinical data and demographics, data on substitute decision-makers and characteristics of the donation consent approach. Study outcomes were consent for organ donation and approach rate. We evaluated independent associations between consent and approach-and system-level factors.Results:We identified 34 837 referrals for organ donation, of which 6548 (18.8%) substitute decision-makers were approached for consent. Of these, 3927 (60.0% of approaches) consented for organ donation and 1883 (48.0% of consents) patients proceeded to be organ donors. The most common reason substitute decision-makers were not approached for consent in a case with donation potential was a late referral by the health care team (45.2%). Modifiable factors independently associated with consent included a telephone approach for consent (adjusted odds ratio [OR] 0.46, 95% confidence interval [CI] 0.35–0.58) and a collaborative approach by a physician and donation coordinator (adjusted OR 1.26, 95% CI 1.01–1.59).Interpretation:Consent for organ donation was associated with several modifiable factors. Organizations should target interventions to ensure timely referrals to organ donation organizations, increase in-person consent approaches and increase physician participation in the approach process.

Many people die on transplant waiting lists because the demand for organs outstrips supply. Almost 4500 people are on organ transplant waiting lists in Canada. Despite public support for organ donation across Canada,¹ donation rates vary between 8.8 and 21.2 donors per million population, ² and a substantial pool of potential donors is not being realized. ^2,3 The identification, referral and approach of potential donors can be facilitated by policy, legislation and best practices, ^3,4 although the efficacy of interventions is variable across jurisdictions.^5,6 Some comprehensive interventions to increase donor numbers have not changed consent rates,⁷ suggesting that the consent approach process may be a target for improvement.Substitute decision-makers play an important role in the organ donation process, even in jurisdictions with donation consent registries or opt-out consent systems. Substitute decision-makers are almost always asked permission for organ donation, even when there is a registered donation consent,⁸ and their consent rates vary widely.⁹ Substitute decision-makers faced with consent decisions often do so in emotionally charged circumstances, and many do not know the explicit wishes of the patient.¹⁰ Given this context, the process of obtaining consent and the supports provided may have a substantial impact on the decision. Practices have been identified that improve consent rates from substitute decision-makers,¹¹ and these are routinely performed by large, high-performing organ donation organizations. Several epidemiological studies have identified nonmodifiable factors associated with donation consent (e.g., race, age, socioeconomic status and education).^12–15 The persistent variability in consent rates suggests that other modifiable factors may influence a substitute decision-maker’s decision to consent.We aimed to identify modifiable approach-and system-level factors that were associated with positive consent for organ donation in Ontario, Canada.     

Yolk sphere formation is initiated in oocytes before development of patency in follicles of the moth,Plodia interpunctella

We describe a provitellogenic stage, a previously unrecognized stage of follicle development in moths, and show that oocytes begin yolk sphere formation prior to the development of patency by the follicular epithelium. The vitellogenic activities of follicles from pharate adult femalePlodia interpunctella (Hübner) were determined by visualizing the subunits of vitellin (YP1 and YP3) and the follicular epithelium yolk protein (YP2 and YP4) using monospecific antisera to each subunit to immunolabel whole-mounted ovaries or ultrathin sections. At 92 h after pupation, yolk spheres that contained only YP2 began to proliferate in the oocytes. The inter-follicular epithelial cell spaces were closed at 92 h making vitellogenin inaccessible to the oocyte, and consequently, the vitellin subunits were not observed in the yolk spheres. YP2 uptake most likely occurred across the brush border from the follicular epithelial cells to the oocyte at this time. At 105 h, the inter-follicular epithelial cell spaces appeared closed yet trace amounts of labeling for vitellin were observed in the spaces and also in the yolk spheres along with YP2. Equivalent labeling for all four YPs in yolk spheres was finally observed at 112 h after pupation when the follicular epithelium had become patent. These data indicate that the provitellogenic stage is an extended transition period between the previtellogenic and vitellogenic stages that lasts for approximately 13 h, and it is marked at the beginning by YP2 yolk sphere formation in the oocyte and at the end by patency in the follicular epithelium.     

Expression and regulation of resistin in osteoblasts and osteoclasts indicate a role in bone metabolism

The adipose tissue is the site of expression and secretion of a range of biologically active proteins, called adipokines, for example, leptin, adiponectin, and resistin. Leptin has previously been shown to be expressed in osteoblasts and to promote bone mineralization, whereas adiponectin expression is enhanced during osteoblast differentiation. In the present study we explored the possible role of resistin in bone metabolism. We found that resistin is expressed in murine preosteoclasts and preosteoblasts (RAW 264.7, MC3T3-E1), in primary human bone marrow stem cells and in mature human osteoblasts. The expression of resistin mRNA in RAW 264.7 was increased during differentiation and seemed to be regulated through PKC- and PKA-dependent mechanisms. Recombinant resistin increased the number of differentiated osteoclasts and stimulated NFkappaB promoter activity, indicating a role in osteoclastogenesis. Resistin also enhanced the proliferation of MC3T3-E1 cells in a PKA and PKC-dependent manner, but only weakly interfered with genes known to be upregulated during differentiation of MC3T3-E1 into osteoblasts. All together, our results indicate that resistin may play a role in bone remodeling.     

Benzoylphenyl ureas inhibit chitin synthesis without interfering with amino sugar uptake in imaginal wing discs of Plodia interpunctella (Hübner)

We tested the hypothesis that the inhibition of chitin synthesis by benzoylphenyl ureas could be explained by their effect on the uptake of GlcNAc into chitin. Our test system consisted of organ cultures of wing imaginal discs from Plodia interpunctella. These wing discs synthesize chitin in response to 20-hydroxyecdysone or RH 5849, a non-steroidal ecdysteroid mimic. Two benzoylphenyl ureas, diflubenzuron and teflubenzuron, inhibited ecdysteroid-dependent chitin synthesis in the wing discs. However, although chitin synthesis was inhibited, there was no corresponding diminution of amino sugar uptake by the imaginal discs. In another experiment 20-hydroxyecdysone stimulated uptake of two sugars, 2-deoxy-D-glucose and 3-O-methyl-D-glucose, which are not synthesized into chitin. Transport of these non-metabolized sugars was unaffected by the inhibitors. In an additional test of the effects on precursor transport, the action of ecdysone (alpha-ecdysone) was examined. Ecdysone stimulated amino sugar uptake, but not chitin synthesis. Neither diflubenzuron nor teflubenzuron inhibited ecdysone-dependent precursor transport. Finally, we examined ecdysteroid-induced uptake of amino sugars by an imaginal disc derived cell line IAL-PID2. In this case, also, GlcNAc transport was not inhibited significantly by either diflubenzuron or teflubenzuron. From these observations we conclude that inhibition of uptake of amino sugars does not account for the ability of teflubenzuron and diflubenzuron to inhibit chitin synthesis in P. interpunctella wing discs.     

Cryptic species,taxonomic inflation,or a bit of both? New species phenomenon in Sri Lanka as suggested by a phylogeny of dwarf geckos (Reptilia,Squamata, Gekkonidae,Cnemaspis)

We evaluated the status of 16 of 22 recognized Sri Lankan Cnemaspis Strauch species, and flagged overlooked diversity with two mitochondrial (cyt b & ND2) and two nuclear markers (RAG1 & PDC) totalling 2829 base pairs. A fossil-calibrated timetree and sampling of other South Asian Cnemaspis provide insights into the diversification of the genus in peninsular India and Sri Lanka. Phylogenetic analyses consistently inferred two broad clades within South Asian Cnemaspis, with Sri Lankan species in two clades, which we call the podihuna and kandiana clades. Each Sri Lankan clade as a whole is sister to Indian taxa and nested within Indian lineages. Cnemaspis modigliani Das from Indonesia is a member of the kandiana clade. This suggests a minimum of two dispersal events between India and Sri Lanka and one between Sri Lanka/India and South-east Asia. South Asian Cnemaspis date back to at least the Eocene, in Sri Lanka to the early Miocene, with late Miocene diversification in the kandiana clade. All but one of the named species we sampled is likely to be valid, and 10 divergent unnamed lineages may warrant specific recognition. A resolution of Sri Lankan Cnemaspis taxonomy will require thorough sampling and the use of both morphological and molecular data.