Establishment of Long-Term Primary Cortical Neuronal Cultures From Neonatal Opossum Monodelphis domestica

Primary dissociated neuronal cultures have become a standard model for studying central nervous system (CNS) development. Such cultures are predominantly prepared from the hippocampus or cortex of rodents (mice and rats), while other mammals are less used. Here, we describe the establishment and extensive characterization of the primary dissociated neuronal cultures derived from the cortex of the gray South American short-tailed opossums, Monodelphis domestica. Opossums are unique in their ability to fully regenerate their CNS after an injury during their early postnatal development. Thus, we used cortex of postnatal day (P) 3–5 opossum to establish long-surviving and nearly pure neuronal cultures, as well as mixed cultures composed of radial glia cells (RGCs) in which their neurogenic and gliogenic potential was confirmed. Both types of cultures can survive for more than 1 month in vitro. We also prepared neuronal cultures from the P16–18 opossum cortex, which were composed of astrocytes and microglia, in addition to neurons. The long-surviving opossum primary dissociated neuronal cultures represent a novel mammalian in vitro platform particularly useful to study CNS development and regeneration.

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Characterization of Postharvest Fungicide-Resistant Botrytis cinerea Isolates From Commercially Stored Apple Fruit

Phytopathology ◽

10.1094/phyto-07-16-0250-r ◽

2017 ◽

Vol 107 (3) ◽

pp. 362-368 ◽

Cited By ~ 16

Author(s):

Wayne M. Jurick ◽

Otilia Macarisin ◽

Verneta L. Gaskins ◽

Eunhee Park ◽

Jiujiang Yu ◽

...

Keyword(s):

Botrytis Cinerea ◽

Gray Mold ◽

Apple Fruit ◽

Sublethal Dose ◽

Long Term Storage ◽

Pose Control ◽

First Time

Botrytis cinerea causes gray mold and is an economically important postharvest pathogen of fruit, vegetables, and ornamentals. Fludioxonil-sensitive B. cinerea isolates were collected in 2011 and 2013 from commercial storage in Pennsylvania. Eight isolates had values for effective concentrations for inhibiting 50% of mycelial growth of 0.0004 to 0.0038 μg/ml for fludioxonil and were dual resistant to pyrimethanil and thiabendazole. Resistance was generated in vitro, following exposure to a sublethal dose of fludioxonil, in seven of eight dual-resistant B. cinerea isolates. Three vigorously growing B. cinerea isolates with multiresistance to postharvest fungicides were further characterized and found to be osmosensitive and retained resistance in the absence of selection pressure. A representative multiresistant B. cinerea strain caused decay on apple fruit treated with postharvest fungicides, which confirmed the in vitro results. The R632I mutation in the Mrr1 gene, associated with fludioxonil resistance in B. cinerea, was not detected in multipostharvest fungicide-resistant B. cinerea isolates, suggesting that the fungus may be using additional mechanisms to mediate resistance. Results from this study show for the first time that B. cinerea with dual resistance to pyrimethanil and thiabendazole can also rapidly develop resistance to fludioxonil, which may pose control challenges in the packinghouse environment and during long-term storage.

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Characterization of cells recovered from the xenotransplanted NG97 human-derived glioma cell line subcultured in a long-term in vitro

BMC Cancer ◽

10.1186/1471-2407-8-291 ◽

2008 ◽

Vol 8 (1) ◽

Cited By ~ 4

Author(s):

Camila ML Machado ◽

Rafael Y Ikemori ◽

Tatiana Q Zorzeto ◽

Ana CMA Nogueira ◽

Suse DS Barbosa ◽

...

Keyword(s):

Cell Line ◽

Glioma Cell ◽

Glioma Cell Line

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Arginine synthesis by hepatomas in vitro. II. Isolation and characterization of Morris hepatoma variants unable to convert ornithine to arginine, and modulation of urea-cycle enzymes by dexamethasone and cyclic-AMP

Journal of Cell Science ◽

10.1242/jcs.68.1.305 ◽

1984 ◽

Vol 68 (1) ◽

pp. 305-319

Author(s):

S.J. Goss

Keyword(s):

Cyclic Amp ◽

Liver Cells ◽

Isolation And Characterization ◽

Rat Liver Cells ◽

Morris Hepatoma ◽

Ornithine Transcarbamoylase ◽

Cellular Incorporation

‘77orn’, a derivative of the Morris rat hepatoma 7777, stably expresses high levels of ornithine transcarbamoylase (OTC) and carbamoylphosphate synthetase I (CPS-I), and is able to grow indefinitely in ornithine-medium (medium with ornithine in place of arginine). Variants that have lost this ability are isolated from 77orn by a ‘suicide’ selective technique dependent on the cellular incorporation of [3H]ornithine. These variants, which have reduced levels of CPS-I, or of both CPS-I and OTC, are shown to have developed multiple hormonal requirements; their enzyme deficiencies can be reversed by use of an appropriately supplemented medium. In particular, CPS-I is inducible by dexamethasone and dibutyryl-cyclic-AMP in combination. Cholera toxin can be used instead of cyclic-AMP, but then butyrate is additionally required if the induction is to be maintained in the long term. The use of these agents in excess can depress OTC. Several other hepatomas, and alos explanted foetal rat liver cells, have similar requirements for CPS-I expression. It is argued that multiple hormonal requirements for CPS-I production are normal in liver cells in vitro, and that hormone-independent hepatomas should be regarded as abnormal. The implications of this for the somatic cell genetic investigation of differentiation are briefly discussed.

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Analysis of in Vitro and in Vivo Characteristics of Human Embryonic Stem Cell-Derived Neural Precursors

Cell Transplantation ◽

10.3727/096368909x484707b ◽

2010 ◽

Vol 19 (4) ◽

pp. 471-486 ◽

Cited By ~ 5

Author(s):

Nataliya Kozubenko ◽

Karolina Turnovcova ◽

Miroslava Kapcalova ◽

Olena Butenko ◽

Miroslava Anderova ◽

...

Keyword(s):

Embryonic Stem ◽

Cellular Heterogeneity ◽

Neural Cells ◽

Lineage Specification ◽

Neural Precursors ◽

Selection Strategies

During the last decade, much progress has been made in developing protocols for the differentiation of human embryonic stem cells (hESCs) into a neural phenotype. The appropriate agent for cell therapy is neural precursors (NPs). Here, we demonstrate the derivation of highly enriched and expandable populations of proliferating NPs from the CCTL14 line of hESCs. These NPs could differentiate in vitro into functionally active neurons, as confirmed by immunohistochemical staining and electrophysiological analysis. Neural cells differentiated in vitro from hESCs exhibit broad cellular heterogeneity with respect to developmental stage and lineage specification. To analyze the population of the derived NPs, we used fluorescence-activated cell sorting (FACS) and characterized the expression of several pluripotent and neural markers, such as Nanog, SSEA-4, SSEA-1, TRA-1-60, CD24, CD133, CD56 (NCAM), β-III-tubulin, NF70, nestin, CD271 (NGFR), CD29, CD73, and CD105 during long-term propagation. The analyzed cells were used for transplantation into the injured rodent brain; the tumorigenicity of the transplanted cells was apparently eliminated following long-term culture. These results complete the characterization of the CCTL14 line of hESCs and provide a framework for developing cell selection strategies for neural cell-based therapies.

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Functional characterization of a stable, noncytolytic stage of macrophage activation in tumors.

Journal of Experimental Medicine ◽

10.1084/jem.146.6.1511 ◽

1977 ◽

Vol 146 (6) ◽

pp. 1511-1520 ◽

Cited By ~ 136

Author(s):

S W Russell ◽

W F Doe ◽

A T McIntosh

Keyword(s):

Macrophage Activation ◽

Functional Characterization ◽

Cytolytic Activity ◽

Target Cells ◽

Tumor Target ◽

Total Absence

The state in which macrophages (Mphi) from regressing Moloney sarcomas could kill tumor target cells was a highly labile one which decayed rapidly in vitro. Thereafter, regressor Mphi were noncytolytic. Mphi from several different progressing sarcomas failed to kill, even when challenged with target cells immediately after explantation. Similarly, thioglycollate-induced peritoneal Mphi (TG-Mphi) did not kill. Noncytolygic Mphi derived either from progressing sarcomas or from long-term (up to 96 h) cultures of regressor Mphi were exquisitely sensitive to stimulation by bacterial lipopolysaccharide (LPS); picogram/milliliter amounts induced killing. Similar concentrations of LPS had no demonstrable effect on TG-Mphi. Thus, tumor Mphi generally appeared to have been primed in vivo, with those in regressing sarcomas having additionally acquired cytolytic activity. Inability of progressor Mphi to kill apparently stemmed from lack of, or failure to respond to, the signal needed in vivo to trigger cytolytic activity, rather than the total absence of activation.

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Characterization of insulin glycation in insulin-secreting cells maintained in tissue culture

Journal of Endocrinology ◽

10.1677/joe.0.1520059 ◽

1997 ◽

Vol 152 (1) ◽

pp. 59-67 ◽

Cited By ~ 31

Author(s):

Y H A Abdel-Wahab ◽

F P M O'Harte ◽

C R Barnett ◽

P R Flatt

Keyword(s):

Tissue Culture ◽

Secretory Activity ◽

Protein Glycation ◽

Subsequent Exposure ◽

Insulin Secreting Cells ◽

Per Se ◽

Stimulation Of

Abstract Characteristics of cellular insulin glycation were examined in the pancreatic B-cell line, BRIN-BD11. The extent of insulin glycation increased stepwise during 72 h of culture at 5·6–33·3 mmol/l glucose, attaining levels up to 27%. Glycation of insulin at 33·3 mmol/l glucose was rapid, reaching maximal values within 2 h, and not readily reversible during 2 to 24 h of subsequent exposure to 5·6 mmol/l glucose. Glycated insulin was readily secreted by BRIN-BD11 cells upon active stimulation with glucose and other secretagogues. Cellular insulin glycation was decreased by 66–80% by inhibitors of protein glycation, vitamin C, aminoguanidine or acetylsalicylic acid. Modulation of insulin-secretory activity of BRIN-BD11 cells by co-culture at high glucose with diazoxide, l-alanine or glibenclamide indicated that long-term stimulation of secretion was associated with a decrease in the extent of insulin glycation. Glycation of insulin in vitro was substantially less extensive than in BRIN-BD11 cells, although glucose-6-phosphate and glyceraldehyde-3-phosphate were 1·4- to 2·0-fold more reactive than glucose per se. These observations indicate that insulin is readily glycated and secreted from insulin-secreting cells under hyperglycaemic conditions in culture. Journal of Endocrinology (1997) 152, 59–67

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Comparative studies ofTrypanosoma (Duttonella) vivaxisolates from Colombia

Parasitology ◽

10.1017/s0031182000074771 ◽

1993 ◽

Vol 106 (1) ◽

pp. 21-29 ◽

Cited By ~ 22

Author(s):

M. F. Dirie ◽

M. J. Otte ◽

R. Thatthi ◽

P. R Gardiner

Keyword(s):

South America ◽

Cross Reactivity ◽

West African ◽

Dairy Calves ◽

South American ◽

Trypanosoma Vivax ◽

Phenotypic Differences ◽

Typical Sign

SUMMARYThe characterization of fourTrypanosoma vivaxisolates from Colombia in South America showed that although minor phenotypic differences existed between them, these parasites are antigenically related and belong to a single serodeme. Characterization by isoenzyme assay, karyotyping and DNA probe analysis, showed the Colombian isolates to be more similar to the West African than to KenyanT. vivax. There was, however, little serological cross-reactivity between South American and African groups ofT. vivax. Although theT. vivaxisolates from Colombia were pathogenic for dairy calves which showed the typical sign of progressive emaciation, these parasites failed to infect mice or tsetseand could not be cultivated as bloodstream formsin vitro.This study represents initial attempts to establish the phenotypic and serological diversity amongstT. vivaxisolates from South America.

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In vitro characterization of commercial factor VIII concentrates: Long-term follow-up

Blut Zeitschrift für die gesamte Blutforschung ◽

10.1007/bf00320384 ◽

1984 ◽

Vol 49 (1) ◽

pp. 53-59 ◽

Cited By ~ 1

Author(s):

C. Miyashita ◽

P. Hellstern ◽

M. K�hler ◽

G. Blohn ◽

E. Wenzel

Keyword(s):

Factor Viii ◽

In Vitro Characterization ◽

Long Term Follow Up ◽

Factor Viii Concentrates

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Comparison of regenerative neurogenesis in response to CNS injury between adult zebrafish and mice

University of Ottawa Science Undergraduate Research Journal ◽

10.18192/osurj.v1i1.3696 ◽

2018 ◽

Vol 1 ◽

pp. 37-41

Author(s):

Kuan-En Chung

Keyword(s):

Radial Glia ◽

Brain Regions ◽

Cns Injury ◽

Adult Zebrafish ◽

Protection Mechanism ◽

Cord Injury ◽

The Difference

The difference between adult zebrafish and mice in their regenerative capacity following central nervous system (CNS) injury is influenced by the permissiveness of the brain microenvironment aside from the intrinsic neurogenic potential of the cell population. In adult zebrafish, glia cells largely retain their radial characteristics and neurogenic capacity, and the zebrafish brain shows full recovery after traumatic brain injury (TBI) as well as spinal cord injury (SCI). Conversely, in mice, radial glia (RG) have largely differentiated into astrocytes. Excluding certain brain regions, following TBI, reactive astrocytes that show the potential to become neural stem cells (NSCs) in vitro remain strictly non-neurogenic in vivo due to the presence of inhibitory factors in the microenvironment. Combined with prolonged inflammation and gliosis, injury to the CNS eventually results in formation of a glial scar further impeding regeneration. However in rodents, suppression of neurogenesis may be a protection mechanism against possible detrimental side-effects of neurogenesis in the long term.

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