Protection against acute paraquat toxicity by ambroxol

An expression vector for G-CSF, pASLB3-3, was constructed and introduced into Namalwa KJM-1 cells (Hosoi et al., 1988), and cells resistant to 100 nM of methotrexate (MTX) were obtained. Among them, the highest producer, clone SC57, was selected and the productivity of this clone was further characterized. The maximal production of G-CSF was at the most 1.8 g/ml/day using a 25 cm² tissue culture flask, even though the cell number was above 7×10⁵ cells/ml. The limiting factors at high density were analyzed as the deficiency of nutrients, such as glucose, cysteine and serine, and pH control. The depression of specific G-CSF productivity per cell under the batch culture conditions was overcome by using a perfusion culture system, Biofermenter^TM (Sato, 1983) with modifications of nutrients supplementation by a dialysis membrane and/or dissolved oxygen (DO) supplementation by microsilicone fibers. ITPSGF medium was modified to elevate concentrations of amino acids and glucose by 2.0- and 2.5-times, respectively. Under the control of pH at 7.4 and DO at 3 ppm, the specific G-CSF productivity was not depressed even at high cell density (above 1×10⁷ cells/ml), and the amount of G-CSF reached 41 g/ml. These results indicated the possibility of finding the optimum culture conditions for the production of recombinant proteins by Namalwa KJM-1 cells.Abbreviations ABTS 2,2-Azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid - BSA Bovine Serum Albumin - BSA-PBS Phosphate-buffered Saline without Ca²⁺ and Mg²⁺ containing Bovine Serum Albumin - dhfr Dihydrofolate Reductase - DO Dissolved Oxygen - G-CSF Granulocyte Colony-stimulating Factor - HEPES 4-(2-Hydroxyethyl)-1-piperazineethansulfonic Acid - IFN Interferon - MTX Methotrexate - PBS(-) Phosphate-buffered saline without Ca²⁺ and Mg²⁺ - Tween-PBS Phosphate-buffered saline without Ca²⁺ and Mg²⁺ containing 0.05% of Tween 20     

Explant organ culture: A review

Organ explant culture models offer several significant advantages for studies of patho-physiologic mechanisms like cell injury, secretion, differentiation and structure development. Organs or small explants/slices can be removed in vivo and maintained in vitro for extended periods of time if careful attention is paid to the media composition, substrate selection, and atmosphere. In the case of human tissues obtained from autopsy or surgery, additional attention must be paid to the postmortem interval, temperature, hydration, and cause of death. Explant organ culture has been effectively utilized to establish outgrowth cell cultures and characterize the histiotypic relationships between the various cell types within an organ or tissue.J. Resau is a visiting scientist at the NCI-LMO-DCE in Frederick, MD 21702, U.S.A.K. Sakamoto is a visiting scientist from the Department of Surgery, Gunma University School of Medicine, Maebashi, Japan     

Passive release of monoclonal antibodies from hybridoma cells

Evidence is presented for the passive release of monoclonal antibodies (MCAB) from hybridoma cells grown in either batch or continuous-flow culture. This release is promoted at room temperature. Passively released MCAB is indistinguishable from that released by actively growing cells, as judged by SDS-polyacrylamide gel electrophoresis. The significance of these observations in relation to the continuous culture of hybridoma cells is discussed.Maximum MCAB content of TB/C3 hybridoma cells is about 55pg per cell, any additional MCAB produced is secreted.Abbreviations MCAB monoclonal antibodies - PBS phosphate buffered saline - RT room temperature - SDS sodium dodecyl sulphate     

Growth of a human yolk sac tumor cell line with yolk sac-derived functions in selenium-supplemented chemically defined synthetic medium

Summary A human yolk sac tumor cell line, TG1, which was established from a testicular yolk sac tumor, was found to replicate continuously in a chemically defined medium supplemented with Na₂SeO₃ (ISRPMI). TG1 produced several plasma proteins and growth factors: albumin, alpha-fetoprotein (AFP), ferritin, carcinoembryonic antigen, beta-2-microglobulin, polyamine, neuron specific enolase, tissue polypeptide antigen, transferrin (Tf), epidermal growth factor, and platelet derived growth factor. By analysis of lectin (LcHA)-affinity electrophoresis, to examine the microheterogeneity of carbohydrate chains of synthetic glycoproteins, TG1 cells cultured with ISRPMI produced only LcHA reactive Tf and AFP based on core fucose attached to asparagine-linkedN-acetylglucosamine residues instead of LcHA-nonreactive Tf and AFP produced by TG1 cells cultured with fetal bovine serum (FBS)-containing medium.α1-6 Fucosyltransferase activity was significantly greater in the TG1 cells cultured with ISRPMI (39.9±1.5 pmol · h⁻¹ · mg⁻¹ protein) than cultured with FBS-containing media (18.2±1.2 pmol · h⁻¹ · mg⁻¹ protein). These results have indicated that the selective increase ofα1-6 fucosyltransferase occurred when the cells were cultured with the FBS-free synthetic media.     

香菇和侧耳属间原生质体的电融合研究

从培养在液体培养基中的香菇、美味侧耳和平菇的单核菌丝用酶法分离了原生质体。施加0.5MHz、500PV/cm的正弦波和μs、6000PV/cm方形脉冲的电场诱导下使其电融合。电融合后的融合子和原生质体在固体培养基上植板培养成菌落。在显微镜下检查融合子菌株菌丝的锁状联合选出从融合子长成的菌株。香菇和美味侧耳的融合菌株产生频率为61.53%,香菇和平菇的融合菌株产生频率为32.58%。根据融合菌株与亲本的拮抗作用和他们的过氧化物同工酶和酯酶同工酶的电泳酶谱与其亲本酶谱的不同,证实这些融合菌株是从融合的异核体生长成的。同时讨论了电融合方法和结果。     

Kinetic studies of phenol degradation by Rhodococcus sp. P1 II. Continuous cultivation

The degradation of phenol by Rhodococcus sp. P1 was studied in continuous culture systems. The organism could be adapted by slowly increasing concentration, step by step, up to 30.0 g · 1^-1 phenol in the influent. The degradation rate reached values of about 0.3 g · g dry mass^-1 ·h^-1. Large step increases in phenol concentration and addition of further substrates (e.g., catechol) were tolerated up to a certain concentration. With increasing dilution rate and increasing inlet phenol concentration the stability of the system decreased.     

Manganese as a controlling factor in mixed cultures of Lactobacillus plantarum and enterobacteriaceae

Addition of manganese, at levels of 50 ppm, to a liquid growth medium simulating adverse silage conditions had no effect on the growth or on the fermentation pattern of Enterobacter cloacae and Proteus vulgaris. Yet, the manganese strongly enhanced the growth of Lactobacillus plantarum. Co-cultures of L. plantarum and E. cloacae or P. vulgaris were, by addition of manganese ions, significantly altered in the favour of the former. This finding can be of use in mixed cultures where Enterobacteriaceae act as spoiler microorganisms.     

Developmental patterns of ornithine decarboxylase activity in organogenesis phase rat embryos in culture and in utero

Summary Ornithine decarboxylase activity was measured during organogenesis in rat embryos grown in utero and whole rat conceptuses maintained in an in vitro culture system. Ornithine decarboxylase levels in vivo showed a distinct peak at embryonic age 10.5 d. Despite identical morphology, protein content, crown rump length and numbers of somites cultured embryos displayed a different developmental pattern and possessed less than half the ODC activity of that in vivo. The data suggest that the normal embryonic programming of ODC activity is significantly altered by the culture environment and that further biochemical comparisons of embryos growing in utero and in vitro may be required to evaluate properly the applicability of this technique to detailed studies of teratogenesis and developmental biology. This work was supported by NIH-5-507-RR5359-17 and a 1980 Research Starter Grant from the Pharmaceutical Manufacturers Association Foundation.     

Multiple Molecular Forms of Acetylcholine Receptors in Cultured Skeletal Muscle Cells: Subcellular Localization and Characterization

Abstract: Skeletal muscle cells of newborn rats, cultured in the absence of neuronal influence, were found to contain two types of cell surface acetylcholine receptors as demonstrated by isoelectric focusing. The isoelectric points of the two types of receptors were indistinguishable from those of junctional and extrajunctional types of receptors in mature animals. The cultured cells had two classes of intracellular α-bungarotoxin (αBT) binding components; one had the same sedimentation coefficient as that of surface receptors (9S), and the other had much smaller apparent molecular weights. Only a single major component was detected by isoelectric focusing analysis of the 9s intracellular aBT binding component, with a PI value close to that of the extra junctional receptor. These results suggest that the junctional and extrajunctional types of receptors may be synthesized through a common precursor.