Decline In Cell Survival Research Articles

Mechanisms regulating the cadmium-mediated suppression of Sp1 transcription factor activity in alveolar epithelial cells

This study demonstrates that in vitro exposure of adult rat alveolar epithelial cells to CdCl 2 decreases DNA binding activity of specificity protein 1 (Sp1), a zinc-finger transcription factor known to play a key role in eukaryotic gene expression, maintenance of homeostasis, cell cycle control, terminal differentiation, and apoptosis. Suppression of Sp1 function, as assessed by electrophoretic mobility shift assays (EMSAs), is dependent upon cadmium (Cd) dose and duration of exposure. A 45% decrease of Sp1 activity occurs as early as 30 min after Cd addition. By 2 h, Sp1 activity is reduced even further with no loss of cell viability, suggesting that Sp1 inactivation precedes cell death. If Cd is removed from cultures during these early periods of exposure, inhibition of Sp1 binding activity is reversed. Sp1 inactivation does not appear to be a generalized, non-selective response to Cd as other transcription factors are up-regulated under the same conditions. Phosphorylation is involved in Sp1 down-regulation, as evidenced by the finding that alkaline phosphatase treatment of nuclear extracts from cells exposed to Cd for 2 h helps restore Sp1 binding activity. A broad spectrum Protein Kinase C (PKC) inhibitor, GF109203X, substantially reduces the Cd-mediated effect on Sp1 suggesting that a member of the PKC family is required for Sp1 phosphorylation. More prolonged Cd exposure promotes Sp1 degradation with the appearance of cleavage products (40 and 50 kDa), as detected by Western blotting. Changes in the integrity of the Sp1 protein are accompanied by a corresponding decline in cell survival. Cd-induced cell death is substantially attenuated if cells are pretreated with antagonists of PKC activity which implies that a PKC isoform is also a participant in this process.

Correlation between cell survival and micronuclei formation in V79 cells treated with vindesine before exposure to different doses of γ-radiation

Effect of 20 nM vindesine sulphate (VDS) treatment was studied on cell survival, growth kinetics and micronuclei induction in V79 cells exposed to 0–300 cGy of γ-radiation at 16, 22 and 28 h post-irradiation. Treatment of V79 cells with VDS before exposure to different doses of gamma radiation resulted in a significant decline in cell survival and growth kinetic when compared with the concurrent PBS+irradiation group. The decline in cell survival and growth kinetics was dose related. Similarly, the cell proliferation indices also declined in a dose dependent manner in both PBS+irradiation and VDS+irradiation groups and this decline was higher in VDS+irradiation group in comparison with the PBS+irradiation group. In contrast, the frequency of micronuclei increased in a dose related manner in both PBS+irradiation and VDS+irradiation groups. However, the frequency of micronuclei was significantly greater in the VDS+irradiation group when compared to the PBS+irradiation group at all the post-irradiation time periods studied and the dose response for both groups was linear for all the scoring time periods. The biological response was determined by plotting surviving fraction and micronuclei frequencies on X- and Y-axes, respectively. The plot between surviving fraction and micronuclei induction showed a close correlation. The surviving fraction of V79 cells reduced with the increasing frequency of micronuclei in both groups and the relationship between micronuclei induction and cell survival could be fitted on a linear quadratic model.

Correlation between cell survival, micronuclei-induction, and LDH activity in V79 cells treated with teniposide (VM-26) before exposure to different doses of γ radiation

The influence of teniposide (VM-26) treatment was studied on the radiation-induced alterations in cell survival, micronuclei (MN) formation and lactate dehydrogenase (LDH) release in V79 cells. Treatment of V79 cells with 10 nM teniposide before exposure to different doses of γ radiation resulted in a significant decline in the cell survival when compared with the PBS+irradiation group. The decline in cell survival was dose related. The cell proliferation indices also declined in a dose-dependent manner in both PBS+irradiation and VM-26+irradiation groups. The decline was higher in the VM-26+irradiaion group in comparison with the PBS+irradiation group. In contrast, the frequency of micronuclei increased in a dose-related manner in both PBS+irradiation and VM-26+irradiation groups. However, the frequency of micronuclei was significantly greater in the latter group when compared with the former group at all the post-irradiation time periods studied. The LDH contents increased in a dose-dependent manner in both PBS+irradiation and VM-26+irradiation groups at all the post-irradiation time periods evaluated. This elevation in LDH contents was significantly greater in the VM-26+irradiation group in comparison with the PBS+irradiation group.

Influence of vindesine exposure on the micronucleus formation and cell survival in V79 cells

Effect of different concentrations (0, 1, 5, 10, 20 and 50 nM) of vindesine sulphate was studied on clonogenicity and micronucleus (MN) formation in V79 (Chinese hamster lung fibroblasts) cells. Exposure of V79 cells to vindesine for 6 h resulted in a concentration dependent decline in cell survival. The frequency of micronuclei (MN) increased in a concentration dependent manner at 16, 22 and 28 h post-exposure. The frequency of MN increased significantly after 5 to 50 nM drug exposure at 16 and 22 h post-treatment, while increasing post-exposure time to 28 h resulted in a significant increase in MN frequency at all exposure doses of vindesine. The statistical evaluation of concurrent concentrations at various time periods showed a non-significant difference in MN frequency among various post-exposure time periods, except 16 h and 28 h for 50 nM, where a significant decline in the MN frequency was observed at 28 h compared to 16 h post-exposure. The cell proliferation indices showed a concentration dependent decline in the frequency of binucleate cells and this decline was linear quadratic. The increasing drug concentration resulted in a concentration dependent decline in cell survival. While the frequency of micronuclei increased the cell survival decreased and the relationship between cell survival and micronucleus induction was linear quadratic.

Osmotic Effects on Ram and Human Sperm Membranes in Relation to Thawing Injury

The effects of hyposmotic and hyperosmotic stresses on ram and human spermatozoa were examined. Human spermatozoa exhibited a precipitate decline in survival at osmolalities below 90 mOsm caused by cells swelling beyond their maximum volume-to-surface area ratio and lysing. Ram spermatozoa exhibited a progressive decline in cell survival at relatively small hyposmotic stresses before exceeding their maximum volume-to-surface area ratio; this prelytic cell loss could be prevented by decreasing the osmolality in a series of 25-mOsm steps. Repeated hyposmotic stress experiments indicated that cells sensitive to prelytic damage constitute a discrete subpopulation within the ram ejaculate. Spermatozoa of both species were apparently resistant to hyperosmotic stresses; human spermatozoa maintained membrane integrity when subject to stresses up to 2.5 Osm and ram spermatozoa up to 1 Osm. However, ram spermatozoa suffered an almost complete and irreversible loss of motility above 600 mOsm. Spermatozoa of both species exposed to hyperosmotic stress and returned to isosmotic conditions exhibited significant cell damage, although ram spermatozoa were the more vulnerable. These observations are related to cryopreservation of spermatozoa.

Activity of phthalocyanine photosensitizers against human glioblastoma in vitro.

In vitro studies utilizing an established human glioblastoma cell line (U87MG) were undertaken to compare the light absorption spectra and photocytotoxicity characteristics of hematoporphyrin derivative (HpD), metal-free phthalocyanine tetrasulfonate (PcS), and aluminum phthalocyanine tetrasulfonate (AlPcS). The peak absorption wavelengths for the compounds studied were: HpD = 630 nm, PcS = 630 nm, AlPcS = 677 nm. Seven different concentrations of each compound (0 to 20 micrograms/ml) were subjected to five different energy levels (40 to 640 joules/cm2) of broad band light (wavelength = 650 +/- 40 nm). Control experiments employing each drug alone and photoactivation alone were also performed. HpD showed a consistent dose-dependent photoactivated cytotoxicity with an abrupt decrease in cell survival when energies of 160 joules/cm2 or more were administered and a drug concentration dependency that resulted in a progressive decline in cell survival beginning at 1.0 microgram/ml. The metal-free PcS had a 10 times greater absorption of light at 675 nm, but only a very weak cytotoxic effect, which occurred at concentrations in excess of 10.0 micrograms/ml and energy levels greater than 320 joules/cm2. AlPcS, however, displayed a 100-fold increase in light absorption at 675 nm when compared to HpD. This was coupled with a decrease in cell survival similar to the HpD curves, but with a much steeper slope of percentage cell kill at lower drug concentrations and energy dosages. The control studies did not result in any appreciable cytotoxic effect. The data suggest that AlPcS holds promise as a second generation photosensitizer in the photodynamic treatment of human gliomas.

The use of survival in aggregation to detect potential carcinogens with retrovirus-infected fischer rat embryo cells

A sensitive and rapid transformation assay procedure is described. The end point, cell survival in aggregation, is measured by counting viable cells dissociated from aggregates in suspension for 4 d. When cells, previously treated with increased concentrations of a carcinogen, are suspended in liquid medium above an agar base, aggregates are formed, while untreated and solvent control cells remain primarily as single cells and show a rapid decline in cell survival; cells that had been treated with chemical carcinogen survive longer in suspension. When used as a marker for in vitro transformation, this end point can assess the carcinogenic potential of an agent.

Chemical enhancement of survival in aggregation of retrovirus-infected rat cells: an interlaboratory comparison.

This study provides a comparative evaluation among three independent laboratories of the responses to 16 chemicals in the retrovirus (Rauscher leukemia) infected Fischer rat embryo (RIFRE) cell-survival-in-aggregation assay. When suspended in liquid media above an agar base, control cells showed a rapid decline in cell survival, whereas cells that had previously been treated with chemical carcinogen survive in suspension longer than control cells. The endpoint, survival in aggregation, is measured by counting viable cells dissociated from aggregates in suspension for 4 days. By modifying previously reported procedures, we have improved the system so that a clear differential (positive or negative) response is achieved by cells treated with either a known carcinogen or known noncarcinogen. Using procedures designed to minimize assay variability, replicate assays were performed and the data analyzed for inter- and intralaboratory concordance. The RIFRE cell-survival-in-aggregation assay demonstrated a high degree of interlaboratory reproducibility in assessing the overall positive or negative responses of known carcinogens and noncarcinogens, and good qualitative reproducibility in assessing compounds tested under code. The assay could discriminate between known carcinogens and noncarcinogens. All chemicals were tested without the addition of a metabolic activation system. Cells exhibiting carcinogen-induced enhancement of survival in aggregation, when plated back onto a solid substrate and carried in culture, subsequently expressed transformation-associated changes in their cellular morphology, growth in semisolid media, and tumorigenicity in nude mice. These results indicate that retrovirus-infected Fischer rat embryo cells detect a carcinogen-mediated early event that progresses to neoplastic phenotypes. Survival in aggregation appears to require the presence of the exogenous retrovirus, since uninfected cells did not show a differential survival response when carcinogen-treated, noncarcinogen-treated, or control cells were compared. This system provides a reproducible method of detecting carcinogenic chemicals based on their ability to induce enhanced survival in aggregation of treated cells.