Enzyme replacement via liposomes variations in lipid composition determine liposomal integrity in biological fluids

Liposomes survive exposure to biological fluids poorly, extruding trapped enzymes, drugs, or solutes upon interaction with serum or plasma constituents. We have quantified the disruptive effects of human serum on liposomes and have studied whether various modifications in their phospholipid composition might produce liposomes with an increased carrier potential for applications in vivo. Multilamellar liposomes (phosphatidylcholine 70:dicetyl phosphate 20: cholesterol 10) were prepared with ³H-labeled phosphatidylcholine as the lipid phase marker and [¹⁴C]inulin and horseradish peroxidase as aqueous phase markers. Gel exclusion chromatography showed that 32 ± 3% of [¹⁴C]inulin and 27 ± 7% of horseradish peroxidase were lost after 1 h incubation with 10% (v/v) human serum. Loss of aqueous solutes was reduced to 20 ± 5%/h and 17 ± 2%/h, respectively, after treatment with decomplemented serum (56°C, 30 min). Loss induced by serum was concentration and time dependent: to 57 ± 2% at 1 h and 67 ± 14% at 24 h, with 50% serum; plasma was slightly less perturbing whereas human serum albumin was not at all disruptive. By incorporating sphingomyelin (35 mol%) into multilamellar liposomes, the leakage of [¹⁴C]-inulin in the presence of 10% serum was reduced to 12 ± 4%/h; increasing the molar percentage of cholesterol to 35% also stabilized the lipid bilayers, reducing leakage to 20 ± 7%/h. Both small and large unilamellar vesicles could not be stabilized against serum-mediated leakage by the incorporation of sphingomyelin. The data suggest that cholesterol and sphingomyelin enhance liposomal integrity in the presence of serum or plasma and promise to yield enhanced survival of drug-laden lipid vesicles in biological fluids in vivo.     

Spray retention for liquid and mycoherbicide inoculum in three weed-biocontrol systems

Spray retention is often used to measure herbicide delivery to optimize application parameters, but little is known about retention characteristics of mycoherbicide inoculum applied for weed biocontrol. This study examined inoculum retention of three mycoherbicide agents, Pyricularia setariae, Colletotrichum truncatum and C. gloeosporioides f. sp. malvae, on their respective weed targets: green foxtail, scentless chamomile and round-leaved mallow. Conidium suspensions of these fungal pathogens containing a sodium-fluorescein tracer dye were applied at 500, 1000 and 2000 L ha^-1 using a cabinet sprayer, and the liquid volume and number of conidia retained on the plants were quantified. On all three weed species, liquid and conidium retention showed a high degree of correlation at varying application volumes although slight differences existed depending on the weed species. Based on the analysis of regression slopes, liquid retention reflected conidium retention most closely on green foxtail but slightly overestimated the number on scentless chamomile and round-leaved mallow, possibly due to different plant morphology and spray run-off at extremely high application volumes. Liquid retention can generally be used as an indicator in studying effects of spray quality on mycoherbicide retention for improved delivery and biocontrol in these weed-biocontrol systems.     

Étude ultrastructurale de la fécondation chez Grantia compressa F.

The fertilization of a calcareous sponge is studied by means of the electron microscope, in Grantia compressa F.. The mature oocyte lies beneath the choanoderm; the carrier cell is applied to its surface, and the spermiocyst enters its cytoplasm. The ultrastructural study demonstrates two formations in this spermiocyst: (i) the ‘head’, surrounded by a lamellar envelope, consists mainly of a nucleus with ring-shaped chromatin; (ii) a large area with a periodic structure corresponding to the modified ‘middle piece’ or ‘mitochondrial body’ of other authors.     

Use of Bottom Ash of Waste Coal as an Effective Microbial Carrier

An experiment was done to determine the efficacy of waste bottom ash as an effective microbial carrier. Bottom ash found to be a suitable microbial carrier. The average of viable cells of Paenibacillus polymyxa GS01 (as a test biocontrol agent) in bottom ash samples was about 10⁸ cfu/10±2 mg. The surface of bottom ash coated with 5% PVA w/v was most effective for improvement of cell viability. TSB medium containing 50 mg/L of MnSO₄·H₂O was the best for spore production of P. polymyxa GS01. Thus waste bottom ash coating with 5% PVA is likely to be suitable for use as a microbial carrier.     

Comparative Proteomic Analysis Identifies Key Metabolic Regulators of Gemcitabine Resistance in Pancreatic Cancer

Pancreatic adenocarcinoma (PDAC) is highly refractory to treatment. Standard-of-care gemcitabine (Gem) provides only modest survival benefits, and development of Gem resistance (GemR) compromises its efficacy. Highly GemR clones of Gem-sensitive MIAPaCa-2 cells were developed to investigate the molecular mechanisms of GemR and implemented global quantitative differential proteomics analysis with a comprehensive, reproducible ion-current–based MS1 workflow to quantify ～6000 proteins in all samples. In GemR clone MIA-GR8, cellular metabolism, proliferation, migration, and ‘drug response’ mechanisms were the predominant biological processes altered, consistent with cell phenotypic alterations in cell cycle and motility. S100 calcium binding protein A4 was the most downregulated protein, as were proteins associated with glycolytic and oxidative energy production. Both responses would reduce tumor proliferation. Upregulation of mesenchymal markers was prominent, and cellular invasiveness increased. Key enzymes in Gem metabolism pathways were altered such that intracellular utilization of Gem would decrease. Ribonucleoside-diphosphate reductase large subunit was the most elevated Gem metabolizing protein, supporting its critical role in GemR. Lower Ribonucleoside-diphosphate reductase large subunit expression is associated with better clinical outcomes in PDAC, and its downregulation paralleled reduced MIAPaCa-2 proliferation and migration and increased Gem sensitivity. Temporal protein-level Gem responses of MIAPaCa-2 versus GemR cell lines (intrinsically GemR PANC-1 and acquired GemR MIA-GR8) implicate adaptive changes in cellular response systems for cell proliferation and drug transport and metabolism, which reduce cytotoxic Gem metabolites, in DNA repair, and additional responses, as key contributors to the complexity of GemR in PDAC. These findings additionally suggest targetable therapeutic vulnerabilities for GemR PDAC patients.     

Urm1 at the crossroad of modifications

The ubiquitin‐like protein Urm1 can be covalently conjugated to other proteins, such as the yeast thioredoxin peroxidase protein Ahp1p, through a mechanism involving the ubiquitin E1‐like enzyme Uba4. Recent findings have revealed a second function of Urm1 as a sulphur carrier in the thiolation of eukaryotic cytoplasmic transfer RNAs (tRNAs). Interestingly, this new role of Urm1 is similar to the sulphur‐carrier activity of its prokaryotic counterparts, strengthening the hypothesis that Urm1 is a molecular fossil of the ubiquitin‐like protein family. Here, we discuss the function of Urm1 in light of its dual role in protein and RNA modification.     

A new type porous carrier and its application to culture of suspension cells

A new type porous carrier was fabricated from a mixture of sodium alginate, bovine serum albumin and sodium bicarbonate. The porous space of the carrier is an assembly of void spaces. The carrier was successfully applied to the cultivation of suspension animal cells. In the culture, while both cells and carriers were held in suspension, the cells were entrapped hydrodynamically into the void spaces in the carriers. A culture of hybridoma cells using this carrier resulted in a cell density up to 5.7×10⁷ cells per ml-carrier.     

Genes regulating copper metabolism

The metabolism of Cu is intimately linked with its nutrition. From gut to enzymes, Cu bioavailability to key enzymes and other components operates through a complex mechanism that uses transport proteins as well as small molecular weight ligands. Steps in Cu transport through the blood, absorption by cells, and incorporation into enzymes are slowly being understood. Cloning and sequencing of the genes for Menkes disease and Wilson disease has shown that membrane-bound enzymes analogous to Cu-ATPases in prokaryotes are equally important to Cu transport and homeostasis in mammalian cells. The primary structure of the mammalian Cu-ATPases has been deduced from cDNAs from tissues and organs. It now appears that mammalian Cu-ATPase have tissue and developmental specificity. In this review, we will focus on the Cu-ATPase that has been identified with Menkes disease. An emphasis will be placed on the existence of multiple forms of the ATPase and some indication as to how the different isoforms befit their role in the normal physiology of copper, specifically transmembrane transport and maintenance of a favorable internal cellular environment.