Fibroblasts, mammalian

In vitro cytotoxicity assays using isolated cells have been applied intermittently to cyanobacterial toxicity testing over several years." Cells investigated for suitability in cyanobacterial toxin assays include primary liver cells (hepatocytes) isolated from rodents and fish, established permanent mammalian cell lines, including hepatocytes, fibroblasts and cancerous cells, and erythrocytes. Earlier work suggested that extracts from toxic cyanobacteria disrupted cells of established lines and erythrocytes," but studies with purified microcystins revealed no alterations in structure or ion transport in fibroblasts or erythrocytes,... 

Jurivich, D. A., Chung, J., Blenis, J. (1991). Heat shock induces two distinct S6 protein kinase activities in quiescent mammalian fibroblasts. J. Cell. Physiol. 148,252-259. 

Key findings that demonstrated that the 0 subunit is the essential component of L-type channels have come from studies of the channel activity of the expressed protein. Expression studies performed in mammalian liver fibroblasts have demonstrated that the oti subunit alone can form a channel [77] and contains the receptors for the DHPs, PAAs and diltiazem [64]. In very elegant studies using a mouse model of muscular dysgenesis it has been demonstrated that the ] subunit DNA can restore Ca currents and the charge movement that arises from the voltagesensing function of the channels to the mutant cells that normally lack these activities [21,78,79]. The restoration of these activities restores excitation-contraction coupling. Thus it is clear that the aj subunit is the major functional unit of L-type Ca channels. 

S-Acylation and Plasma Membrane Targeting of the Farnesylated Carboxyl-Terminal Peptide of N-Ras in Mammalian Fibroblasts, H. Schroeder, R. Leventis, S. Rex, M. Schelhaas, E. Nagele, H. Waldmann, J. R. Silvius, Biochemistry 1997, 36,13102-13109. 

S. cerevisiae aurora/IPllp, C. elegans aurora/AIR-2, and mammalian aurora B (also called AIM-1) phosphorylate H3 at Ser-10 and Ser-28 during mitosis [39 1] (Fig. 5). Protein phosphatase 1 removes the phosphate at these sites [39,42]. INCENP is bound to aurora B and is essential for the proper targeting of aurora B on the chromosomes [43-45]. INCENP and aurora B (AIM-1) are overexpressed in a variety of human cancers, including colorectal cancer [43,46]. Overexpression of aurora B (AIM-1) in CHE diploid fibroblasts leads to chromosomal instability, suggesting that aurora B overexpression may play a role in carcinogenesis [46]. INCENP/aurora B and H3 phosphorylation appear to be involved in assembly of mitotic chromosomes, but not mitotic chromosome compaction [44,47]. 

Since the brain expresses many mRNAs that are also found in non-nervous tissue and are therefore of little interest to the psychopharmacologist, it is necessary to isolate only those cDNAs that, for example, encode for a specific enzyme or receptor protein. Several techniques have been developed to achieve this. For example, a specific cDNA plasmid may be inserted into cultured mammalian cells such as fibroblasts that can express the specific receptor or enzyme. Once this has been expressed in the culture medium, the receptor or enzyme can be identified by adding a specific ligand or substrate. This enables those cells that expressed the specific macromolecule of interest to be identified and... 

Specific cDNAs (for example a receptor) inserted in plasmid that transfects a mammalian cell (e.g. fibroblasts) in culture... 

Cytotoxicity assays employ mammalian celk in culture to measure cellular metabolic impairment and death resulting from exposure in vitro to soluble and particulate toxicants. Mammalian cells derived from various tissues and organs can be maintained as short term primary cultures or, in some cases, as continuous cell strains or lines. The cytotoxicity assays, incorporated as part of Level 1 analysis, employ primary cultures of rabbit alveolar (lung) macrophages (RAM) and maintenance cultures of strain WI-38 human lung fibroblasts. 

A variety of cell lines, strains, or primary cell cultures, including human cells, may be used (e.g., Chinese hamster fibroblasts, human or other mammalian peripheral blood lymphocytes). Cell cultures are exposed to the test substance both with and without metabolic activation and at predetermined intervals after exposure, they are treated with a metaphase-arresting substance (e.g., colchicine), harvested, stained, and metaphase cells are analyzed microscopically for the presence of structural chromosome aberrations. At least three concentrations should be used. 

Mammalian cells in culture are exposed to the test substance. Established cell lines are treated both with and without metabolic activation. Cells are incubated for an appropriate length of time, then rinsed, fixed, and dried. Slides are developed, stained, and exposed silver grains are counted. The endpoint of UDS is measured by determining the uptake of labeled nucleosides in cells that are not undergoing scheduled (S-phase) DNA synthesis. The most widely used technique is the determination of the uptake of H-TdR by autoradiography. Primary cultures (e.g., rat hepatocytes), human lymphocytes, or established cell lines (e.g., human diploid fibroblasts) may be used in the assay. Multiple concentrations of the test substance over a range adequate to define the response, should be used. 

Resorcinol was not genotoxic in bacterial assays or in in vivo mammalian assays it did cause chromosomal aberrations in human lym-phoqn es in vitro but not in cultured human fibroblasts. ... 

Circadian oscillation profiles of clock genes are induced in several mammalian peripheral culture cells by serum shock (Balsalobre et al 1998). To elucidate whether serum induces the circadian expression of human clock genes in normal human diploid fibroblasts, we applied RT-PCR ELISA methods to detect RNA levels of clock genes in WI-38 cells after serum stimulation (Fig. 3). Since WI-38 cells in culture invariably undergo senescence after a finite number of doublings, we selected young WI-38 cells. The RT-PCR-ELISA data are expressed as amounts (Moles) of corresponding cDNA plasmids in... 

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