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PubMedDate not supplied

[Rehydration conditions for dehydrated Saccharomyces cerevisiae yeasts and cell viability].

The viability of dehydrated Saccharomyces cerevisiae populations was studied as a result of changes in the conditions of rehydration (temperature, pH, time, composition of the medium). Rehydration was shown to play a key role for the functional characteristics of populations in the general consideration of dehydration-rehydration and to involve several factors. The viability of cells is regarded as both a relative and absolute criterion for the physiological state of populations. The conditions of rehydration should be strictly standardized for dehydrated microorganisms in comprehensive studies of mechanisms involved in the stability of microorganisms.

Culture Media↗

PubMed2004

Sam35 of the mitochondrial protein sorting and assembly machinery is a peripheral outer membrane protein essential for cell viability.

The mitochondrial outer membrane contains two integral proteins essential for cell viability, Tom40 of the translocase of the outer membrane (TOM complex) and Sam50 of the sorting and assembly machinery (SAM complex). Here we report the identification of Sam35, the first peripheral mitochondrial outer membrane protein that is essential for cell viability. Sam35 (encoded by the Saccharomyces cerevisiae ORF YHR083w) is a novel subunit of the SAM complex and is crucial for the assembly pathway of outer membrane beta-barrel proteins, such as the precursors of Tom40 and porin. Sam35 is not required for the import of inner membrane or matrix targeted proteins. The presence of two essential proteins in the SAM complex, Sam35 and Sam50, indicates that it plays a central role in mitochondrial biogenesis.

Amino Acid Sequence↗

PubMed2003

[Comparative assessment of the human cell viability ex vivo in Hanks solutions in distilled and Penta water].

Assessment of human colon and lung mucosa cell viability was performed in Hanks salt media prepared separately with distilled and patented Penta water. The cell viability in the suspension was estimated by fluorescence intensity of propidium iodide, a DNA specific dye, that is an indicator of DNA structure intactness or damage. The experiments were conducted with the flow cytometry technique. The histogram analysis showed that 2-hour incubation of the cells in Hanks salt medium prepared with distilled water resulted in an increase of the number of the apoptotic cells with a respective decrease of the number of the intact cells (approximately 2- and 4-fold in the suspensions of the colon and lung mucosa cells respectively). A similar experiment with Hanks salt medium prepared with Penta water resulted in a less marked increase of the viability of the apoptotic cells that did not exceed 20 and 50% for the colon and lung mucosa cells respectively. The findings showed that viability of the cells ex vivo was significantly higher when Penta water was used as a solvent for preparing Hanks salt media as compared to distilled water. The result is important for ex vivo experiments since maximum preservation of the DNA structure minimizes the number of possible experimental inaccurate and consequently erroneous conclusions. Furthermore, the fact of pathologic process inhibition in cells isolated from various human tissues in Penta-based salt media is in favour of using Penta water as a solvent for nutritional ingredients in ex vivo maintenance of human tissues for transplantation as compared to distilled water.

Cell Survival↗

PubMed1995

Heat shock protein 70 overexpression affects the response to ultraviolet light in murine fibroblasts. Evidence for increased cell viability and suppression of cytokine release.

To elucidate cellular concepts for protection against ultraviolet (UV) light we investigated the effect of heat shock protein 70 (hsp70) overexpression on cell viability and on the secretion of UV-inducible immunological cytokines. Transfected murine fibrosarcoma cells (WEHI-S), overexpressing hsp70 or a sham transfected control were used. Overexpression of hsp70 was sufficient to markedly increase cell viability upon treatment with UVB (290-320 nm). Since long wave UV (UVA, 320-400 nm) as well as UVB turned out to stimulate the release of O2- radicals we studied the cell viability upon oxidative stress. Hsp70 overexpression increased viability upon treatment with hydrogen peroxide or menadione, but had no influence on UV-induced O2- release. UV-light is known to upregulate immunologic and proinflammatory cytokines such as IL-1 and IL-6. Oxidative stress appeared to exert a similar effect. Hsp70 overexpression markedly decreased the release of IL-6 induced by UVA, UVB and oxidative stress. To test whether the hsp70 mediated suppression is confined to events caused by UV-light we determined IL-1-mediated effects. IL-1-induced IL-6 release was reduced by hsp70 overexpression, whereas the IL-1 mediated activation of nuclear factor kappa B was not affected. Our data suggests that hsp70 plays a central role not only in cell protection against UV-light, but also in the regulation of proinflammatory cytokine release induced by UV-exposure.

Animals↗

PubMedDate not supplied

Photo-cross-linking of type I collagen gels in the presence of smooth muscle cells: mechanical properties, cell viability, and function.

The effectiveness of photomediated cross-linking of type I collagen gels in the presence of rat aortic smooth muscle cells (RASMC) as a method to enhance gel mechanical properties while retaining native collagen triple helical structure and maintaining high cell viability was investigated. Collagen was chemically modified to incorporate an acrylate moiety. Collagen methacrylamide was cast into gels in the presence of a photoinitiator along with RASMC. The gels were cross-linked using visible light irradiation. Neither acrylate modification nor the cross-linking reaction altered collagen triple helical content. The cross-linking reaction, however, moved the denaturation temperature beyond the physiologic range. A twelve-fold increase in shear modulus was observed after cross-linking. Cell viability in the range of 70% (n = 4, p > 0.05) was observed in the photo-cross-linked gels. Moreover the cells were able to contract the cross-linked gel in a manner commensurate with that observed for natural type I collagen. Methacrylate-mediated photo-cross-linking is a facile route to improve mechanical properties of collagen gels in the presence of cells while maintaining high cell viability. This enhances the potential for type I collagen gels to be used as scaffolds for tissue engineering.

Acrylamides↗

PubMed1999

Coexpression of CD9 augments the ability of membrane-bound heparin-binding epidermal growth factor-like growth factor (proHB-EGF) to preserve renal epithelial cell viability.

BACKGROUND: Transfection of renal epithelial cells (NRK 52E) with membrane-associated heparin-binding epidermal growth factor-like growth factor (proHB-EGF) increased renal epithelial cell survival by promoting cell-cell and cell-extracellular matrix interactions. ProHB-EGF has been shown to form a complex in the plasma membrane with the tetraspanin CD9, an interaction that significantly increases the effectiveness of proHB-EGF as a juxtacrine mitogenic agent. METHODS: We examined whether the coexpression of proHB-EGF and CD9 would increase renal epithelial cell survival. CD9 was stably transfected into NRK 52E cells, either alone (NRKCD9) or together with proHB-EGF (NRKboth). RESULTS: Juxtacrine mitogenic activity of NRKCD9 was no different than in cells transfected with vector alone (NRKvector), but was increased by NRKboth; juxtacrine mitogenic activity by NRKboth was twofold greater than when proHB-EGF was transfected alone (NRKproHB-EGF). When grown in 10% fetal calf serum, growth rates were similar among all transfectants. However, in 1% fetal calf serum, NRKproHB-EGF grew 50% faster than NRKvector or NRKCD9, and NRKboth grew 20% to 50% faster than NRKproHB-EGF at one, two, and three days of culture. NRKproHB-EGF attachment to plastic substratum at one, two, and three hours was 250% greater than that of NRKvector, and NRKboth was 20% to 30% greater than that of NRKproHB-EGF. Coating plates with either poly 2-hydroxyethyl methacrylate or the GRGDTP peptide prevented normal cell-extracellular matrix attachment, and NRKvector or NRKCD9 failed to attach or form cell-cell attachments. NRKproHB-EGF exhibited 300% and NRKboth exhibited 600% greater cell viability under these conditions. Expression of type I and type III collagen mRNA was enhanced similarly in NRKproHB-EGF and NRKboth, but the expression of beta1 integrin was up-regulated only in NRKboth. CONCLUSIONS: Coexpression of proHB-EGF and CD9 may render the renal epithelial cells more resistant to disruption of cell-cell and cell-matrix interactions and could accelerate the re-establishment of these attachments.

Animals↗

PubMed1992

Improvement of endothelial cell viability at 4 degrees C by addition of lazaroid U74500A to preservation solutions.

Lazaroids are potent inhibitors of lipid peroxidation. Endothelial cell damage has been shown to occur during cold storage preservation of lung and liver. This study examines the effects on endothelial cell viability of the addition of four lazaroids, U74006F, U78518F, U74500A, or U75412E to preservation solutions. Human umbilical vein endothelial cell cultures were stored at 4 degrees C for 48 and 96 hr in EuroCollins or 5% polyethylene glycol in buffered saline (PEG). U78518F, U74500A, U74006F, U75412E, or dexamethasone (each 50 microM) was added to EC (n = 32) or PEG (n = 32) and compared with control solutions of EC or PEG alone. Endothelial cell viability was determined by measuring cellular reduction of 3-[4,5-dimethylthiazol-2-yl]-2,3-diphenyltetrazolium bromide to a purple formazan dye. The reduction occurs only in viable cells and requires mitochondrial dehydrogenase activity. Results were quantified by measuring dye absorbance (Ab) with a micro-ELISA spectrophotometer. Absorbance values were compared by ANOVA and reported as mean values +/- standard deviation. Addition of U74500A to EC (Ab = 0.474 +/- 0.055) and PEG (Ab = 0.462 +/- .005) improved viability at 48 hr when compared with EC (Ab = 0.289 +/- 0.069) and PEG (Ab = 0.287 +/- 0.052) alone (P less than 0.05). At 96 hr, addition of U74500A resulted in improved viability in both EC (Ab = 0.377 +/- 0.068) and PEG (Ab = 0.195 +/- 0.09) or PEG alone (Ab = 0.212 +/- 0.1) (P less than 0.05). Other lazaroids tested were also effective in improving cellular viability, but to a lesser degree than U74500A. This study demonstrates that the addition of lazaroids to organ preservation solutions improves endothelial cell viability.

Absorption↗

PubMed2004

EGCG down-regulates telomerase in human breast carcinoma MCF-7 cells, leading to suppression of cell viability and induction of apoptosis.

Telomerase is elevated in >90% of breast carcinomas and therefore has received much attention as a target for breast cancer therapy and cancer diagnostic research. Dietary components that are capable of inhibiting the growth of cancer cells without affecting the growth of normal cells are receiving considerable attention in developing novel cancer-preventive approaches. Studies have shown that (-)-epigallocatechin-3-gallate (EGCG) from green tea imparts a growth inhibitory effect on cancer cells. Here, we show that treatment of EGCG dose-dependently inhibited (20-100%) the reproductive or colony forming potential, and also decreased cell viability at different time points studied ( approximately 80% inhibition) in human breast carcinoma MCF-7 cells but had no adverse effect on the growth of normal mammary cells. Treatment of EGCG for 48 and 72 h markedly increased the percentage of apoptotic cells (32-51%) in MCF-7 cells compared to that of non-EGCG treated cells (8-14%). In order to identify the possible mechanism of decreased cell viability and induction of apoptosis in breast carcinoma cells by EGCG, we found that treatment of MCF-7 cells with EGCG dose-dependently inhibited telomerase activity (40-55%), and also inhibited the mRNA expression (40-55%) of hTERT, a catalytic subunit of telomerase. Additional studies demonstrated that EGCG also inhibited the protein expression of hTERT, which indicated that inhibition of telomerase was associated with down-regulation of hTERT. Together, our results indicate that EGCG down-regulates telomerase in human breast carcinoma MCF-7 cells, leading to the suppression of cell viability and induction of apoptosis, thus providing the molecular basis for the development of EGCG as a novel chemopreventive and pharmacologically safe agent against breast cancer.

Antineoplastic Agents, Phytogenic↗

PubMed2002

Blockade of TGF-beta inhibits mammary tumor cell viability, migration, and metastases.

TGF-betas are potent inhibitors of epithelial cell proliferation. However, in established carcinomas, autocrine/paracrine TGF-beta interactions can enhance tumor cell viability and progression. Thus, we studied the effect of a soluble Fc:TGF-beta type II receptor fusion protein (Fc:TbetaRII) on transgenic and transplantable models of breast cancer metastases. Systemic administration of Fc:TbetaRII did not alter primary mammary tumor latency in MMTV-Polyomavirus middle T antigen transgenic mice. However, Fc:TbetaRII increased apoptosis in primary tumors, while reducing tumor cell motility, intravasation, and lung metastases. These effects correlated with inhibition of Akt activity and FKHRL1 phosphorylation. Fc:TbetaRII also inhibited metastases from transplanted 4T1 and EMT-6 mammary tumors in syngeneic BALB/c mice. Tumor microvessel density in a mouse dorsal skin window chamber was unaffected by Fc:TbetaRII. Therefore, blockade of TGF-beta signaling may reduce tumor cell viability and migratory potential and represents a testable therapeutic approach against metastatic carcinomas.

Animals↗

PubMed1997

Correlation of discocyte frequency and ATP concentration in preserved blood. A morphological indicator of red blood cell viability.

Red blood cells (RBC) are viable if kept in an adequate preservative solution, although gradual changes in morphology and metabolism may occur. There is a gradual decrease in adenosine-5'-triphosphate (ATP) concentration, pH, glucose consumption, and enzyme activity during preservation. The normal discocyte shapes are initially replaced by echinocytes and stomatocytes and, at final stages, by spherocytes, the last step before splenic sequestration. Post-transfusional survival has been correlated with the ATP concentration. RBC preserved in ADSOL, a solution containing adenine, dextrose, sodium chloride, and mannitol, are viable for transfusion for up to 6 weeks. Erythrocytes from 10 blood units taken from healthy adult donors were preserved for 12 weeks in ADSOL at 4 degrees C. We now report a significant correlation (r2 = 0.98) between the percentage of discocytes (89 to 7%) and ATP (100 to 10%) concentration in ADSOL-preserved RBC. The results suggest that the percent of discocyte shapes used as an indicator of ATP concentration may be a useful indicator for quality control of RBC viability in centers which have limited assay facilities.

Adenosine Triphosphate↗

PubMed2000

Novel biphasic effect of pyrrolidine dithiocarbamate on neuronal cell viability is mediated by the differential regulation of intracellular zinc and copper ion levels, NF-kappaB, and MAP kinases.

Nuclear factor kappaB (NF-kappaB) is a transcription factor involved in the expression of a wide range of genes, most of which code for proteins that play a role in immunity and inflammation. Pyrrolidine dithiocarbamate (PDTC) is a well-known inhibitor of NF-kappaB. Although its mechanism of action is conferred by its antioxidant property, other mechanisms by which PDTC can act as a prooxidant, metal chelator, and free thiol group modulator have recently been suggested. Here we report that PDTC caused a dual effect on cell viability in neuronal rat pheochromocytoma (PC12) cells, depending on its concentration. Increase of intracellular zinc and copper ion levels selectively potentiated the cytotoxic PDTC effect in a dose-dependent manner, and thiol reagents, such as glutathione and N-acetylcysteine, as well as divalent metal-chelating reagents, such as EDTA and bathocuproline disulfonic acid, blocked its cell death effect. The differential effect of PDTC on cell viability correlates well with the inhibition of NF-kappaB activities. In addition, PDTC differentially activated microtubule-associated protein (MAP) kinases, such as extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), but not p38, depending on its dose, and the coaddition of glutathione (GSH), other antioxidants, and metal ions also modulated their activities. Furthermore, stable Bcl-2 expression blocked the PDTC-induced cell death. These results suggest that the thiol groups and free zinc and copper ion levels are important for the novel biphasic PDTC effect on cell viability, which is associated with the differential activation of NF-kappaB and MAP kinases.

Animals↗

PubMed1999

Effects of Ca(2+)-ionophore A23187 and calmodulin antagonists on regulatory mechanisms of glycolysis and cell viability of NIH-3T3 fibroblasts.

We studied here, in NIH-3T3 fibroblasts, the effect of the Ca(2+)-ionophore A23187 (which is known to increase intracellular-free Ca(2+)) on the control of glycolysis and cell viability and the action of calmodulin antagonists. Time-response studies with Ca(2+)-ionophore A23187 have revealed dual effects on the distribution of phosphofructokinase (PFK) (EC 2.7.1.11), the rate-limiting enzyme of glycolysis, between the cytoskeletal and cytosolic (soluble) fractions of the cell. A short incubation (maximal effect after 7 min) caused an increase in cytoskeleton-bound PFK with a corresponding decrease in soluble activity. This leads to an enhancement of cytoskeletal glycolysis. A longer incubation with Ca(2+)-ionophore caused a reduction in both cytoskeletal and cytosolic PFK and cell death. Both the "physiological" and "pathological" phases of the Ca(2+)-induced changes in the distribution of PFK were prevented by treatment with three structurally different calmodulin antagonists, thioridazine, an antipsychotic phenothiazine, clotrimazole, from the group of antifungal azole derivatives that were recently recognized as calmodulin antagonists, and CGS 9343B, a more selective inhibitor of calmodulin activity. The longer incubation with Ca(2+)-ionophore also induced a decrease in the levels of glucose 1,6-bisphosphate and fructose 1,6-bisphosphate, the two allosteric stimulatory signal molecules of glycolysis. All these pathological changes preceded the reduction in cell viability, and a strong correlation was found between the fall in ATP and cell death. All three calmodulin antagonists prevented the pathological reduction in the levels of the allosteric effectors, ATP and cell viability. These experiments may throw light on the mechanisms underlying the therapeutic action of calmodulin antagonists that we previously found in treatment of the proliferating melanoma cells, on the one hand, and skin injuries, on the other hand.

3T3 Cells↗

PubMed2005

Interaction of CmeABC and CmeDEF in conferring antimicrobial resistance and maintaining cell viability in Campylobacter jejuni.

OBJECTIVES: To determine the role of CmeDEF in conferring antimicrobial resistance in Campylobacter jejuni and examine the interaction of CmeABC and CmeDEF in mediating antimicrobial resistance and maintaining cell viability. METHODS: Single and double mutants of cmeF and cmeB were generated in multiple strains using insertional mutagenesis. The mutants were compared with their wild-type strains for antimicrobial susceptibility and growth characteristics. Transcription fusion was used to quantify the expression of cmeDEF and cmeABC. Ethidium bromide (EB) accumulation assay was used to measure the efflux function. RESULTS: Insertional mutagenesis of the cmeF gene in C. jejuni NCTC 11168 resulted in a 2-fold decrease in the resistance to ampicillin, polymyxin B and EB, whereas the same mutation in C. jejuni 81-176 and 21190 led to a 2-4-fold increase in the resistance to multiple antimicrobials and toxic compounds. The increased resistance in the cmeF mutants of 81-176 and 21190 was associated with the elevated efflux in the mutants. Compared with the cmeB mutant, the cmeF/cmeB double mutants of 81-176 and 21190 showed further decrease in the resistance to various antimicrobials and toxic compounds. Transcription fusion assay indicated that the expression level of cmeF was substantially lower than that of cmeB. Notably, the cmeB/cmeF double mutation, not the single mutations, impaired cell viability in Campylobacter. CONCLUSIONS: CmeDEF interacts with CmeABC in conferring antimicrobial resistance and maintaining cell viability in C. jejuni. CmeABC is the predominant efflux pump in C. jejuni, whereas CmeDEF plays a secondary role in conferring intrinsic resistance to antimicrobials.

Anti-Bacterial Agents↗

PubMed1994

Interrelationship of changes in islet nicotine adeninedinucleotide, insulin secretion, and cell viability induced by interleukin-1 beta.

Complete loss of pancreatic insulin function in insulin-dependent diabetes is thought to be due to an autoimmune cytokine-mediated destruction of the beta-cell. The effects of several classes of agents on interleukin-1 beta (IL-1 beta)-induced suppression of insulin secretion, beta-cell NAD levels, and beta-cell viability were examined. After overnight incubation of isolated rat islets with 15 U/ml IL-1 beta and 11 mM glucose, sequential hourly insulin secretory responses to the same glucose concentration, 22 mM glucose, and 22 mM glucose plus forskolin were severely inhibited to 10-37% of the control value. Islet NAD levels were also sharply reduced to 43% of the control value after 24-h exposure to IL-1 beta, but not after 1 or 3 h, demonstrating the same time course as that for inhibition of insulin secretion. Exposure to IL-1 beta also decreased islet cell viability measured as trypan blue exclusion. Only 1 mM N-methyl arginine, an inhibitor of nitric oxide synthase, completely protected all three parameters of beta-cell function from damage by IL-1 beta. Nicotinamide and thymidine prevented the IL-1 beta-induced loss of cell viability and suppression of NAD, but had no effect on sustaining insulin secretion. Antioxidants, steroids, and several neuropeptides also did not prevent inhibition or restore the secretory response. Thus, the loss of the secretory response appears to be more narrowly restricted to nitric oxide radical damage induced by exposure to IL-1B.

Amino Acid Oxidoreductases↗

PubMed2002

Single-cell viability assessment with a novel spectro-imaging system.

Single-cell viability assessment by means of plural dye probes require the spectral and temporal analysis of microscopic images of the test cells. To meet this requirement, we have developed a simple and compact spectro-imaging system using an image slicer and a grism. The image slicer was made of a bundle of 100 optical fibers. The field of view is divided into 10 x 10 sections. The spectral data of each section could be recorded every 5 s in the range from 400 to 800 nm at 5 nm resolution. The viability changes of yeast or tobacco single-cells were measured with this system. Using BY-2 cells, for example, the response to a chemical stress of saponin was measured by means of two fluorescent probes. The spectral-spatial-temporal data of fluorescein and DNA bound ethidium bromide provided us with useful information about the dynamic change of cell membrane permeability from which the cell viability was assessed.

Amphotericin B↗

PubMed2001

MRI study of immediate cell viability in focused ultrasound lesions in the rabbit brain.

The purpose of this study was to evaluate cell viability in MR imaged focused ultrasound (FUS) lesions using cell-viability staining with triphenyl tetrazolium chloride (TTC) and both light and electron microscopy. Ten paired ultrasonic lesions were created in 5 rabbit brains in vivo with an ultrasound beam of 1.5 MHz electrical power input to the transducer of 50 W and exposure duration of 15 seconds. T2-weighted fast spin-echo (FSE) MRI was performed to detect the FUS lesions in the brain 4 hours after treatment, after which the animals were immediately euthanized. Lesion sizes were measured on TTC-stained specimens, histological sections stained with hematoxylin and eosin (H&E), and T2-weighted MR images. The differences between the lesion diameters measured with the three methods were within the range of 0.1--0.7 mm. The lesion sizes measured from MRI correlated well with those seen from H&E sections. The measurements from MRI slightly overestimated lesion sizes on TTC-stained wet tissues by approximately one MRI pixel (0.31 mm). Electron microscopy demonstrated nuclear and cytoplasmic ultrastructural damage within the grey-white, non-TTC-stained lesion zone, whereas the TTC-stained normal tissue showed preservation of neuronal ultrastructure. Therefore, MR-imaged lesions represent a cell-death zone in rabbit brain 4 hours after FUS ablation, with slight overestimation by approximately one MRI pixel. J. Magn. Reson. Imaging 2001;13:23-30.

Animals↗

PubMed2003

Human hepatocyte isolation and relationship of cell viability to early graft function.

Hepatocyte transplantation is emerging as an additional modality of treatment for patients with acute liver failure or liver-based metabolic disorders. The procedure requires isolation of high-quality hepatocytes from unused donor livers. Hepatocytes were isolated from 20 donor livers (11 right lobes, 3 left lateral segments, 6 whole livers) using a collagenase perfusion technique. Cell viability (median 56%, range 13-95%) and yield (median 1.4 x 10(9) cells, range 2.0 x 10(6)-1.8 x 10(10) cells) varied according to the tissue available. Fatty livers rejected for transplantation gave lower cell viability (median 45%, range 25-59%). There was a significant correlation between age of donor (median 21 years, range 7-66 years) and viability of isolated hepatocytes in vitro (r = -0.683, p = 0.001). The 13 segments of livers were from reduced/split grafts used for clinical transplantation in 9 children and 4 adults. There was no significant correlation between in vitro cell viability and clinical parameters including intensive care stay, serum aspartate aminotransferase,and international normalized ratio (in the first 7 days), and allograft rejection or other early posttransplant complications, in patients transplanted with the corresponding tissue.

Adolescent↗

PubMedDate not supplied

Influence of mechanical stress on cell viability.

The cartilage is a hydrated connective tissue in joints that withstands and distributes mechanical forces. The chondrocytes utilize mechanical signals to regulate their metabolic activity through complex biological and biophysical interactions with the extracellular matrix (ECM). The aim of this work was to study the influence of mechanical stress on cells behavior cultured in 3D biosystems (alginate and alginate supplemented with hyaluronate). After mechanical stimulation, cell viability and cell death process were the main studied parameters. Our results indicated that viability and cell cycle progression were inhibited under mechanical stimulation, as far as the extracellular matrix was not yet synthesized. In contrast, on day 21, the mechanical stimulation had positive effect on these parameters.

Alginates↗