Treatment with Extracellular Vesicles from Giardia lamblia Alleviates Dextran Sulfate Sodium-Induced Colitis in C57BL/6 Mice

Inflammatory bowel disease (IBD) is a chronic and recurrent illness of the gastrointestinal tract. Treatment of IBD traditionally involves the use of aminosalicylic acid and steroids, while these drugs has been associated with untoward effects and refractoriness. The absence of effective treatment regimen against IBD has led to the exploration of new targets. Parasites are promising as an alternative therapy for IBD. Recent studies have highlighted the use of parasite-derived substances, such as excretory secretory products, extracellular vesicles (EVs), and exosomes, for the treatment of IBD. In this report, we examined whether EVs secreted by Giardia lamblia could prevent colitis in a mouse model. G. lamblia EVs (GlEVs) were prepared from in vitro cultures of Giardia trophozoites. Clinical signs, microscopic colon tissue inflammation, and cytokine expression levels were detected to assess the effect of GlEV treatment on dextran sulfate sodium (DSS)-induced experimental murine colitis. The administration of GlEVs prior to DSS challenge reduced the expression levels of pro-inflammatory cytokines, including tumor necrosis factor alpha, interleukin 1 beta, and interferon gamma. Our results indicate that GlEV can exert preventive effects and possess therapeutic properties against DSS-induced colitis.     

The molecular weight and concentration of dextran sulfate affect cell growth and antibody production in CHO cell cultures

To investigate the effect of dextran sulfate (DS), a widely used anti‐aggregation agent, on cell growth and monoclonal antibody (mAb) production including the quality attributes, DS with the three different MWs (4,000 Da, 15,000 Da, and 40,000 Da) at various concentrations (up to 1 g/L) was added to suspension cultures of two different recombinant CHO (rCHO) cell lines producing mAb, SM‐0.025 and CS13‐1.00. For both cell lines, the addition of DS, regardless of the MW and concentration of DS used, improved cell growth and viability in the decline phase of growth. However, it increased mAb production only in the CS13‐1.00 cells. Among the three different MWs, 40,000 Da DS was most effective in attenuating cell aggregation during the cultures of CS13‐1.00 cells, and showed the highest maximum mAb concentration. For SM‐0.025 cells, it significantly decreased specific mAb productivity, particularly at a high concentration of DS. Overall, DS addition did not negatively affect the quality attributes of mAbs (aggregation, charge variation, and glycosylation), though its efficacy on mAb quality depended on the MW and concentration of DS and cell lines. For both cell lines, the addition of DS did not affect N‐glycosylation of mAbs and decreased basic charge variants in mAbs. For CS13‐1.00 cells, the mAb monomer increased with the addition of 40,000 Da DS at 0.3–1.0 g/L. Taken together, to maximize the beneficial effect of DS addition on mAb production, the optimal MW and concentration of DS should be determined for each specific rCHO cell line. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1113–1122, 2016     

The effects of apolipoprotein B depletion on HDL subspecies composition and function

HDL cholesterol (HDL-C) efflux function may be a more robust biomarker of coronary artery disease risk than HDL-C. To study HDL function, apoB-containing lipoproteins are precipitated from serum. Whether apoB precipitation affects HDL subspecies composition and function has not been thoroughly investigated. We studied the effects of four common apoB precipitation methods [polyethylene glycol (PEG), dextran sulfate/magnesium chloride (MgCl₂), heparin sodium/manganese chloride (MnCl₂), and LipoSep immunoprecipitation (IP)] on HDL subspecies composition, apolipoproteins, and function (cholesterol efflux and reduction of LDL oxidation). PEG dramatically shifted the size distribution of HDL and apolipoproteins (assessed by two independent methods), while leaving substantial amounts of reagent in the sample. PEG also changed the distribution of cholesterol efflux and LDL oxidation across size fractions, but not overall efflux across the HDL range. Dextran sulfate/MgCl₂, heparin sodium/MnCl₂, and LipoSep IP did not change the size distribution of HDL subspecies, but altered the quantity of a subset of apolipoproteins. Thus, each of the apoB precipitation methods affected HDL composition and/or size distribution. We conclude that careful evaluation is needed when selecting apoB depletion methods for existing and future bioassays of HDL function.     

Transformation of Glycyrrhizic Acid by Aspergillus spp.

Glycryrrhizic acid was metabolized to 3-oxo-18β-glycyrrhetinic acid via 18β-glycyrrhetinic acid by Aspergillus niger, A. oryzae, A. sojae, and A. tamarii. Two methyl esters were derived from these two metabolites and identified by their ¹³C-NMR spectra and MS data.     

The effects of ruthenium red,lanthanum, fluorescein isothiocyanate and trifluoperazine on vesicle transport,vesicle fusion and tip extension in pollen tubes

The effects of ruthenium red, lanthanum, fluorescein isothiocyanate and trifluoperazine, all antagonists of Ca²⁺ function in cells, have been studied in growing pollen tubes of Tradescantia virginiana. All four drugs inhibit pollen-tube growth but bring about different ultrastructural changes at the growing tips and within the cytoplasm. The results strongly support the hypothesis that Ca²⁺ plays a vital role in the mechanism of pollen-tube tip growth. The effect of ruthenium red provides evidence that sequestration of Ca²⁺ by mitochondria critically adjusts the concentration of these ions at tube tips. Fluorescein isothiocyanate appears to be a potent inhibitor of vesicle fusion at the plasma membrane, with vesicles accumulating in the tip at rates equivalent to those determined previously for their production. Both vesicle fusion and tip extension are regulated by Ca²⁺ but appear to be independently controlled processes.     

Growth performances ofLactobacillus hilgardii immobilized in dextran gel and in continuous fermentation

A variant ofLactobacillus hilgardii was immobilized by its own production of dextran gel, forming grains. The best rate of weight increase of the gel in continuous fermentation was 16.3±3.3%/h, at pH 4.8±0.1 and with a dilution rate of 0.22 to 0.26/h. Observation by scanning electron microscopy located most of the bacteria as microcolonies on the surface. A similar arrangement appeared in calcium alginate beads. The best population density (10¹⁰ cells/g) was obtained in grains at pH 5.8, after 30h. At a similar pH value, 4.8, the growth rate was higher in alginate beads than in dextran gel but the final population density was approximately the same. Acidification rate increased faster with mixed gel at pH 5.2 than with dextran at pH 5.8.     

The potential for heparin and its derivatives in the therapy and prevention of HIV-1 infection

Heparin is one of several sulphated polysaccharides which potently inhibit replication of the human immunodeficiency virus type 1 (HIV-1) in cultures of CD4;ve human cells. The EC50 value is around 5 μg ml-1. We have demonstrated that heparin binds to recombinant gp120, the envelope glycoprotein of HIV-1, at a site termed the V3 loop, or principle neutralizing domain, which consists of a disulphide-bridged loop of 32–35 amino acids particularly enriched with basic residues. Using a series of chemically modified heparins we have shown that there is structural specificity in the anti-HIV activity of heparin. Heparin is routinely used clinically as an anticoagulant, and has proved essentially non-toxic and well tolerated. Low anticoagulant derivatives of heparin which retain high anti-HIV-1 activities in vitro may be generated by several routes. Such preparations are ideal candidates for clinical investigation as potential novel therapeutic agents for use in combination with other drugs in the management of AIDS and HIV infection. This revised version was published online in November 2006 with corrections to the Cover Date.     

Leishmania donovani: Therapeutic and prophylactic action of antimony dextran glycoside (RL-712) in the golden hamster

Antimony dextran glycoside (RL-712, manufactured by Rosco A/S Pharmaceutical Taastrup, Denmark) produced a remarkable decrease in Leishmania donovani densities in hamster spleen and liver when administered in a single intraperitoneal (ip) dose of 500 mg Sb/kg body weight or in 600 mg Sb/kg body weight divided into two or three equal parts injected at weekly intervals beginning 15 days after intracardial inoculation of 10 million amastigotes, and also in a single ip dose of 300 mg Sb/kg body weight 14, 7, and 0 days before intracardial inoculation of 10 million amastigotes. No parasites were detected on using a single dose of 500 mg Sb/kg body weight followed by 100 mg Sb/kg body weight one week later.When the drug was administered in a dose of 500 mg Sb/kg body weight either 10 days before subcutaneous inoculation of 60 million promastigotes or eight days before intracardial inoculation of five million promastigotes, no parasites were recovered by smear or culture from the spleen and liver.Results show the prophylactic and therapeutic actions of this drug against L. donovani in the hamster.     

Estimating viability of plant protoplasts using double and single staining

Summary The utility of numerous dyes for determining the viability of barley (Hordeum vulgare L. cv. Himalaya) aleurone protoplasts was studied. Protoplasts isolated from the barley aleurone layer synthesize and secrete -amylase isozymes in response to treatment with gibberellic acid (GA) and Ca²⁺. These cells also undergo dramatic morphological changes which eventually result in cell death. To monitor the viability of protoplasts during incubation in GA and Ca²⁺, several types of fluorescent and nonfluorescent dyes were tested. Evans blue and methylene blue were selected as nonfluorescent dyes. Living cells exclude Evans blue, but dead cells and cell debris stain blue. Both living and dead cells take up methylene blue, but living cells reduce the dye to its colorless form whereas dead cells and cell debris stain blue. The relatively low extinction coefficient of these dyes sometimes makes it difficult to distinguish blue-stained cells against a background of blue dye. Several types of fluorescent dyes were tested for their ability to differentially stain dead or living cells. Tinopal CBS-X, for example, stains only dead cells, and its high extinction coefficient allows its ultraviolet fluorescence to be recorded even when preparations are simultaneously illuminated with visible light. To double-stain protoplasts, the most effective stain was a combination of fluorescein diacetate (FDA) and propidium iodide (PI). By employing a double-exposure method to record the fluorescence from cells stained with both FDA and PI, dead and living cells could be distinguished on the basis of fluorochromasia.     

Elevation of rat intestinal permeability by enterotoxigenic Escherichia coli in comparison to non-toxigenic E. coli and Salmonella typhimurium. Application of fluorescent dextran 3000 as permeability probe

Abstract The effect of bacterial enterotoxins on rat intestinal permeability properties was studied by comparing the effect of toxin-positive and toxin-negative Escherichia coli and Salmonella typhimurium inoculated into a segment of rat small intestine. Fluoresceinated dextran 3000 (FITC-D3; M _r 3000) was applied as permeability marker. The E. coli strain C922a-1 producing heat-labile (LT) and heat-stable (ST) enterotoxins and colonising factor CFA/II increased the transmural passage of the dextran probe into portal blood. In contrast, its plasmid-negative variant, a non-toxin producer lacking CFA, caused permeability changes indistinguishable from the bacteria-free nutrient broth control. Another pair of enterotoxigenic E. coli strains, 1628–14 (LT⁺, ST⁺, CFA/I⁺) and 1628–15 (LT⁺, ST⁻ and CFA/I⁻) both increased the intestinal permeability. The observations indicate that the LT⁺-only E. coli strain 1628–15 has the ability to promote permeability of rat intestine. The toxin-negative, rough S. typhimurium 395MR10 bacteria had a very small effect on the permeability, which was also achieved with culture filtrate only.
It is concluded that enterotoxigenic E. coli (ETEC) can alter the properties of the mucosal barrier towards intermediate-sized molecules that could be of antigenic significance, or which could play a crucial role in the nutritional status of the host organism.