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PubMed2006

Rapid test for distinguishing membrane-active antibacterial agents.

In the search for antibacterial agents with a novel mode-of-action (MOA) many targeted cellular and cell-free assays are developed and used to screen chemical and natural product libraries. Frequently, hits identified by the primary screens include compounds with nonspecific activities that can affect the integrity and function of bacterial membrane. For a rapid dereplication of membrane-active compounds, a simple method was established using a commercially available Live/Dead(R) Bacterial Viability Kit. This method utilized two fluorescent nucleic acid stains, SYTO9 (stains all cells green) and propidium iodide (stains cells with damaged membrane red) for the drug-treated bacterial cells. The cells were then either examined visually by fluorescence microscopy or their fluorescence emissions were recorded using a multi-label plate reader set to measure emissions at two different wavelengths. The ratio of green versus red was compared to a standard curve indicating the percentage of live versus dead bacteria. Nine known antibiotics and 14 lead compounds from various antibacterial screens were tested with results consistent with their MOA.

Anti-Bacterial Agents↗
PubMed2005

Using fluorescence for improvement of the quantitative analysis of micronucleus in cell culture.

The micronucleus test is of unquestionable importance, being widely used in studies with several purposes. Standardization of the technique has been proposed by several groups mainly when in vivo assays are performed. In cell cultures the staining method that predominates is the Feulgen's reaction, since it is specific for DNA. In this work, we evaluated the use of two fluorescent stains, SYTOX green and propidium iodide, in substitution for the Feulgen's reaction. Based on the results we concluded that the fluorescence microscopy allows the reliable detection of micronuclei, presenting the following advantages: the technique is easy and fast to perform, it avoids acid treatments; the fluorescence is long lasting; micronucleus counting becomes easier and more reliable by the use of RNAase treatment; the technique is more sensitive to detect the smallest micronuclei.

Animals↗
PubMed2005

Isatin, an endogenous monoamine oxidase inhibitor, triggers a dose- and time-dependent switch from apoptosis to necrosis in human neuroblastoma cells.

Isatin is an endogenous indole that is increased in stress, inhibits monoamine oxidase (MAO) B and improves bradykinesia and striatal dopamine levels in rat models of Parkinson's disease. Consequently, it has been suggested that isatin might be a possible treatment for Parkinson's disease although little is known about its effects on neural cell growth and survival. The aim of this study was to investigate the survival of dopaminergic human neuroblastoma (SH-SY5Y) cells following treatment with increasing concentrations of isatin. SH-SY5Y cells were exposed to isatin for defined time points, after which cell survival was determined by MTT assay. A combination of Annexin V binding and propidium iodide (PI) exclusion was used to distinguish apoptosis from necrosis in flow cytometry experiments and FACS profiles of permeabilised PI-labelled cells were employed for the assessment of cell cycle distribution. Isatin treatment (1-400 microM) for 24h induced a significant dose-dependent increase in MTT metabolism by SH-SY5Y cells in culture, but this was not due to an increase in cell division. At the higher concentrations (200-400 microm) isatin triggered cell death, although MTT metabolism was still increased in the culture, suggesting that surviving cells were hypermetabolic. Following a longer (48 h) exposure, isatin was found to cause cell death in a dose-dependent manner; at lower concentrations (50 microM), the predominant mode of cell death was apoptosis while at the highest concentration (400 microm) increasing numbers of necrotic cells were also evident. Thus, in dopaminergic SH-SY5Y cells isatin induces cell death in dose- and time-dependent manner. This death occurred as a continuum of survival, apoptosis and necrosis. Our results re-emphasise that caution should be exercised when considering high doses of isatin as a putative anti-Parkinson's disease therapeutic.

Annexin A5↗
PubMed2004

Ibuprofen and apigenin induce apoptosis and cell cycle arrest in activated microglia.

In case of injury or disease, microglia are recruited to the site of the pathology and become activated as evidenced by morphological changes and expression of pro-inflammatory cytokines. Evidence suggests that microglia proliferate by cell division to create gliosis at the site of pathological conditions such as the amyloid plaques in Alzheimer's disease and the substantia nigra of Parkinson's disease patients. The hyperactivation of microglia contributes to neurotoxicity. In the present study we tested the hypothesis that anti-inflammatory compounds modulate the progression of cell cycle and induce apoptosis of the activated cells. We investigated the effects of ibuprofen (non-steroidal anti-inflammatory drug) and apigenin (a flavonoid with anti-inflammatory and anti-proliferative properties) on the cell cycle of the murine microglial cell line BV-2. The findings indicate that apigenin-induced cell cycle arrest preferentially in the G2/M phase and ibuprofen caused S phase arrest. The binding of annexin V-FITC to the membranes of cells which indicates the apoptotic process were examined, whereas the DNA was stained with propidium iodide. Both apigenin and ibuprofen induced apoptosis significantly in early and late stages. The induction of apoptosis by ibuprofen and apigenin was confirmed using TUNEL assay, revealing that 25 microM apigenin and 250 microM ibuprofen significantly increased apoptosis in BV-2 cells. The results from the present study suggest that anti-inflammatory compounds might inhibit microglial proliferation by modulating the cell cycle progression and apoptosis.

Animals↗
PubMed2006

Voltage-dependent potassium channels are involved in glutamate-induced apoptosis of rat hippocampal neurons.

The role of voltage-dependent potassium channel currents in glutamate-treated rat hippocampal neurons was investigated. Cell viability was evaluated by MTT reduction assay and morphological changes. Apoptosis was detected by Hoechst33342 staining with fluorescent microscopy and propidium iodide staining with flow cytometry. Membrane potassium channel currents were recorded with whole-cell patch clamp recordings. Results showed that after shortly exposed to glutamate, about 25 and 50% cells died in 3 h and 24 h, respectively. Meanwhile, the enhancement of IK was observed within 6 h after the glutamate insult. TEA selectively blocked IK and significantly reduced cell apoptosis. IA did not change in the insult though 4-AP, the blocker of this current, showed a protective effect against the injury. These data were in consistent with the hypothesis that K+ efflux contributed to glutamate-triggered apoptosis and IK channels might have a therapeutic effect on the treatment of cerebral ischemia.

Animals↗
PubMed2004

S-methylcysteine may be a causal factor in monohalomethane neurotoxicity.

S-methylcysteine (SMC) is formed after exposure to monohalomethanes in rodents as well as in humans. The present study was performed to study whether SMC, directly or indirectly, contributes to the well-known neurotoxicity of monohalomethanes. We have investigated the effects of acute exposure to SMC by means of electrophysiolocal measurements in freshly prepared hippocampal slices and dissociated hippocampal neurons in culture. For longer-term exposures (24 h) we have used organotypic cultures (2 weeks in culture), taking electrophysiologic recordings and assessing membrane integrity with propidium iodide (PI) fluorescence. We found that only high concentrations of SMC (10(-2) M; exposure time 30 min) in freshly isolated slices of adult rats reduce synaptically evoked population spikes in the CA1 region. This effect was at least partially reversible. In organotypic cultures, at 5 x 10(-5) M after 24 h of exposure, SMC compromises membrane integrity as revealed by PI fluorescence, only in the dentate gyrus, spreading to pyramidal cell layers at 50 x 10(-4) M. At 5 x 10(-6) and 2 x 10(-5) M, under the same experimental conditions, no changes were seen with the PI method, but we recorded increased population spike amplitudes, repetitive discharges and frequency potentiation (at a stimulus repetition rate of 0.05 Hz). Using whole-cell patch clamp in hippocampal dissociated neurons we have found that SMC (applied for approximately 1s) reduces GABA-induced currents ( IC(50) = 4.4 x 10(-4) M) without having an effect of its own, acting like a competitive antagonist at GABA(A) receptors. Our findings are in line with the view that the ability of monohalomethanes to induce the formation of SMC is an important factor for their neurotoxicity, provided that SMC is allowed to act at least for several hours. The effects exerted by SMC seem to be due, at least in part, to its interaction with GABA receptors.

Animals↗
PubMed2006

Cytotoxic effects of various stressors on PC12 cells: involvement of oxidative stress and effect of antioxidants.

In order to specifically elucidate the involvement of oxidative stress, the effects of various types of stressors and antioxidants on PC12 cells were examined. In this study, the following four stressors were studied in detail: free radicals generated from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH), 4-hydroxynonenal (HNE), 7-ketocholesterol (KC), and arsenic trioxide (As2O3). Undifferentiated PC12 cells were treated with 50% lethal concentration (LC50) of these stressors, and subsequently the viability, apoptosis/necrosis ratio, reactive oxygen species (ROS) production, caspase-3 activity, and protection by antioxidants were measured to elucidate the underlying mechanisms that determine the action of these stressors on PC12 cells. The cytotoxicity did not correlate directly with the intracellular formation of ROS. For example, as compared to AAPH, As2O3 produced considerably smaller amounts of ROS at LC50. As observed in the cells incubated with As2O3, KC and HNE exerted cell toxicity, but with a moderate production of ROS. With the exception of HNE, the apoptosis/necrosis ratio of all the stressors evaluated by annexin V and propidium iodide assays increased with an increase in the incubation time at the LC50 values of these stressors. In accordance with apoptosis ratio, caspase activity was detected in the cells incubated with AAPH, As2O3, and KC, but not HNE at LC50 for 24 h. The protective effect of alpha-tocopherol, 17beta-estradiol, 2,3-dihydro-5-hydroxy-2,2-dipentyl-4,6-di-tert-butylbenzofuran (BO653), glutathione, and N-acetylcysteine (NAC) against cytotoxicity depended on the type of stressors. These antioxidants were found to be effective against the abovementioned stressors, except As2O3 against which only NAC was effective. These results suggest that the involvement of ROS and the protective effect of antioxidants depend on the type of stressors.

Aldehydes↗
PubMed2006

Irbesartan attenuates contrast media-induced NRK-52E cells apoptosis.

BACKGROUND: Radiocontrast nephropathy (RCN) is a major complication after radiographic examination. The precise mechanisms underlying RCN are not well understood. Renal tubular cell apoptosis is a feature of RCN, but hyperosmolality cannot fully explain the cytotoxicity of contrast media. There is accumulating evidence that reactive oxygen species (ROS) is involved in the pathophysiology of RCN, whereas the correlation between oxidative stress and contrast media-induced cell apoptosis is not clear. We hypothesized that ROS mediated apoptosis in renal tubular cells exposed to contrast media. Irbesartan, a selective AT(1) receptor antagonist has been demonstrated an antioxidative effect. The present study was designed to determine whether irbesartan attenuated the contrast media-induced renal tubular cell apoptosis. METHODS: NRK-52E cells were exposed to increasing concentration (25, 50, 100, 150 mgiodinemL(-1), 335, 384, 420, 521 mOsmkg(-1)) of ioversol (a non-ionic contrast media) for 1h or incubated in ioversol (100 mgiodinemL(-1), 420 mOsmkg(-1)) for 15 min, 30 min, 60 min, 120 min, 240 min, respectively. Mannitol with the same osmolality as ioversol (420 mOsmkg(-1)) also treated NRK-52E cells for 1h. In separate experiment, irbesartan (0.01, 0.1, 1 mmolL(-1)) was added 1h before incubation with ioversol (100 mgiodinemL(-1), 420 mOsmkg(-1)) for 1h. Apoptosis was determined by Hoechst staining and flow cytometry with annexinV-FITC and propidium iodide. The intracellular formation of ROS was detected by confocal microscopy with fluorescent probe CM-H2DCFDA. Bax and bcl-2 mRNA expression were determined by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Ioversol induced NRK-52E cells apoptosis in a concentration- and time-dependent manner. The intracellular ROS generation was greatly increased following ioversol stimulus. Furthermore, ioversol induced a decrease in the expression for bcl-2 mRNA and an increase for bax mRNA. Irbesartan attenuated the ioversol-induced apoptosis in NRK-52E cells in a dose-dependent manner, in which the protective effect of irbesartan was dependent on decreasing intracellular ROS formation. In addition, irbesartan reversed the ioversol-induced increase in bax mRNA and decrease in bcl-2 mRNA. CONCLUSION: Ioversol induced NRK-52E cells apoptosis in a concentration- and time-dependant manner via an increase in oxidative stress and subsequent to the increase in mRNA expression for bax and reduction in bcl-2 mRNA. Irbesartan attenuated the ioversol-induced apoptosis in NRK-52E cells by reducing oxidative stress and reversing the enhancement of bax mRNA and the reduction in bcl-2 mRNA.

Animals↗
PubMedDate not supplied

Morphological and intracellular alterations induced by Serratia marcescens cytotoxin.

In the present work, in vitro assays were used to investigate the toxicity of Serratia marcescens cytotoxin in cultured Chinese hamster ovary (CHO) cells. The time necessary to detect cellular alterations such as the onset of apoptosis, the perturbation of mitochondrial function, and cytoskeletal changes was assessed. The internalization of the cytotoxin by CHO cells was also examined. Within 10-15 min of exposure to cytotoxin, CHO cells became round, the nucleus shrank, the chromatin became more compact, and cytoplasmic blebs appeared on the cell surface. TUNEL (TdT-mediated dUTP nick end labeling) and propidium iodide staining identified some nuclei with fragmented DNA, and electrophoresis of CHO cell DNA obtained after 30-min exposure to S. marcescens toxin showed a pattern of DNA fragments typically associated with apoptosis. The cells also lost their characteristic actin organization within 10 min of exposure to cytotoxin. Lactate dehydrogenase leakage was detected after 20-min exposure to the cytotoxin and increased with time thereafter. Concomitantly, there was a time-dependent reduction in mitochondrial activity. Fluorescein-labeled S. marcescens cytotoxin was detected only on the surface of CHO cells, even after 30-min exposure to the toxin. These results show that there was no internalization of the toxin by CHO cells, and that, once bound to the cell surface, the toxin was able to induce changes in intracellular metabolism and to trigger cell death by apoptosis.

Animals↗
PubMed2003

Lovastatin-induced cardiac toxicity involves both oncotic and apoptotic cell death with the apoptotic component blunted by both caspase-2 and caspase-3 inhibitors.

The objective of this study was to evaluate the cardiac toxicity of the HMG-CoA reductase inhibitors by testing the hypothesis that lovastatin induces apoptotic and/or oncotic cell death in the myocyte element of the heart and further that cell death is mediated through interruption of the mevalonate pathway and that apoptosis is induced through activation of caspase-2 and caspase-3. Cardiomyocytes were cultured from embryonic chick heart. Lovastatin-induced apoptosis in these cells was demonstrated by three independent techniques, namely (1) FACS analysis of low DNA content by propidium iodide (PI); (2) microscopic assessment for cellular changes of apoptosis; and (3) FACS analysis of cells stained with PI and fluorescein diacetate. Lovastatin produced a concentration-dependent increase in apoptotic cell death and 100 microM lovastatin showed over a 4-fold increase in apoptosis compared to control. Lovastatin also induced oncotic cell death, as there was a 2.5-fold increase in the amount of oncotic cell death compared to control. Lovastatin-induced apoptosis operated, in part, through the mevalonate pathway. The caspase-2 inhibitor z-VDVAD-fmk and the caspase-3 inhibitor Ac-DEVD-CHO reduced the extent of lovastatin-induced cardiac apoptosis. In contrast, lovastatin-induced oncosis was not only insensitive to these caspase-2 or -3 inhibitors but occurred through a mevalonate-independent mechanism of action. In summary, lovastatin-induced cardiotoxicity is complex and represents the sum of two distinct modes of cell death operating in part through the mevalonate pathway with the apoptotic component subject to modification by inhibitors of the initiator caspase, caspase-2, as well as the effector caspase, caspase-3.

Animals↗
PubMed2004

Effect of human chorionic gonadotrophin supplementation during different culture periods on in vitro maturation of canine oocytes.

The IVM of canine oocytes is characterized by low rates of metaphase II. The objective of this study was to evaluate the effects of hCG on meiotic development of canine oocytes for culture periods up to 96 h. Oocytes were collected after ovariohysterectomy. Only oocytes >110 microm in diameter, with a homogeneous dark cytoplasm and three or more layers of compact cumulus cells were used. For IVM, the COCs were cultured in TCM-199+10% fetal calf serum, without (medium A control) or supplement with 10 IU/mL of hCG (medium B), or with a combination of both media (treatment B/A). The COCs were randomly allocated into three groups. The first and second groups were cultured in either medium A or B, respectively for 24, 48, 72, and 96 h. Oocytes of the third group (treatment B/A) were incubated in medium with hCG (medium B) the first 48 h and then transferred to medium without hCG (medium A) for an additional 24 or 48 h. The proportion of COCs with cumulus cell expansion was also evaluated before fixation. Oocytes were stained with propidium iodide prior to nuclear assessment (with epifluorescence microscopy). COCs with cumulus expansion were evident after 48 h of culture. The proportion of COCs with cumulus expansion was higher (P<0.05) for media containing hCG (B or B/A) than for meda lacking hCG (A); this difference was maintained for 72 and 96 h in culture. In media A, B and B/A, 23.3, 31.7 and 29.5%, respectively, of oocytes were at metaphase II after 72 h, with 20.7, 33.1 and 43.4% at this stage after 96 h. The advancement of meiosis was directly proportional to the time of incubation; the highest percentage (P<0.05) of oocytes at metaphase II was observed after 96 h of culture when 10 IU/mL hCG was present for only the first 48 h of culture.

Animals↗
PubMed2004

Relationship between non-return rate and chromatin condensation of deep frozen bull spermatozoa.

This study analyzes the relationship between chromatin condensation and field fertility, expressed as 90 days non-return rate (NRR), of bulls actively used by AI studs. Frozen-thawed semen from five bulls (six ejaculates per bull, three straws per ejaculate), that showed a non-return rate between 60 and 80%, were analyzed to assess sperm chromatin condensation and stability. The chromatin condensation was determined by flow cytometry using propidium iodide as fluorochrome, and the chromatin stability was evaluated by inducing its decondensation with SDS and EDTA. Coefficient of variation among replicates was less than 7% and 5% for chromatin condensation and stability, respectively. No correlation was present between chromatin condensation and NRR. However, significant correlation was found between chromatin stability and NRR. Chromatin stability was higher (P < 0.05) in those bulls that showed higher fertility. The results obtained in this study conclude that assessment of stability could be a valuable tool for routine evaluation and identification of ejaculates with high levels of sperm chromatin abnormalities and to detect animals of higher reproductive potential.

Animals↗
PubMed2007

Dialysis of boar semen prior to freezing-thawing: its effects on post-thaw sperm characteristics.

Low-molecular weight components of the seminal plasma have a detrimental effect on sperm function. The present study was undertaken to evaluate the effect of the removal of low-molecular weight components by dialysis on sperm characteristics prior to and after freezing. Semen, collected from 5 boars, was extended in Kortowo-3 extender (K-3, Poland) and cooled for 3h (control non-dialysis) or dialyzed for 5h in semi-permeable dialysis bags of 12-14kDa molecular weight cut-off prior to freezing. The semen samples were diluted in lactose-hen egg yolk-glycerol extender (lactose-HEY-G) or lactose-lyophilized lipoprotein fractions-glycerol extender (lactose-LPFo-G), packaged into aluminum tubes and frozen in a controlled programmable freezer. Pre-frozen and frozen-thawed spermatozoa were evaluated for motility, plasma membrane (SYBR-14 and propidium iodide) and acrosome integrity, mitochondrial function (Rhodamine 123) and ATP content. The results of the study showed that dialysis significantly improved the sperm characteristics prior to freezing. Dialysis enhanced (P<0.05) post-thaw sperm motility, plasma membrane integrity and mitochondrial function, but had no significant effect (P>0.05) on recovery of spermatozoa with intact acrosomes. Furthermore, dialyzed spermatozoa exhibited higher (P<0.05) ATP content compared with the control after freezing-thawing. Consistent inter-boar variability was detected mainly in dialyzed semen following freezing-thawing. These results indicated that the improvement in sperm quality characteristics prior to freezing and the post-thaw sperm recovery were due to the removal of low-molecular weight components from the seminal plasma. It can be suggested that dialysis is effective in improving the post-thaw quality of boar spermatozoa and has also great practical importance in improving the protocols for cryopreservation of semen. Dialysis may also contribute to a better understanding of different mechanisms underlying cryo-induced damage to boar spermatozoa.

Acrosome↗
PubMed2004

Tri-n-butyltin-induced cytotoxicity on rat thymocytes in presence and absence of serum.

Influence of fetal bovine serum (FBS) on cytotoxicity induced by tri-n-butyltin (TBT), an environmental pollutant, on rat thymocytes was examined to reveal how FBS modifies TBT cytotoxicity. As the medium FBS concentration was increased from 0 to 10%, the cytotoxicity of TBT was dose-dependently reduced when the cells were incubated with 1 microM TBT for 3 h. Almost complete inhibitions of TBT-induced changes in cell viability and population of cells with exposed phosphatidylserine (cells undergoing apoptosis) were observed when the FBS concentration was 10%. Thus, the cytotoxicity induced by 3 h incubation with TBT in FBS-free medium may be different from that in medium containing 10% FBS. However, even in presence of 10% FBS, TBT at concentrations ranging from 10 to 300 nM exerted cytotoxic action on rat thymocytes when the cells were incubated with TBT for 24 h. TBT dose-dependently increased the population of shrunken cells, of which more than 30% were stained with propidium. TBT at 30 nM or more significantly increased the population of cells with hypodiploid DNA, indicating TBT-induced apoptotic cell death. Thus, in the presence of 10% FBS, the prolonged incubation (24 h) of rat thymocytes with TBT at nanomolar concentrations induced apoptosis rather than necrosis.

Animals↗
PubMed2006

Different effects of two cyclic chalcone analogues on cell cycle of Jurkat T cells.

It has been previously shown that the cyclic chalcone analogues E-2-(4'-methoxybenzylidene)- (2) and E-2-(4'-methylbenzylidene)-1-benzusuberone (3) inhibited proliferation of various murine and human tumor cells. In order to gain new insights into the cytotoxic mechanism of the two compounds detection of apoptosis and necrosis of Jurkat T cells exposed to 2 and 3 were performed by flow cytometry using the Annexin V-FITC and propidium iodide double staining method. Analysis of the DNA histograms at 8, 24 and 48 h exposure times showed that near equitoxic doses of 2 and 3 had different effects on the cell cycle of the exposed cells. The immediate (8h) effect of 2 was a remarkable decrease of cells in the G(0)/G(1) phase and increase in the G(2)/M phase. This effect could also be seen in the histogram of cells at the 24h time point. On the contrary, such an effect of 3 could not be observed. At the 24 and 48 h time points accumulation of sub-G(0) (apoptotic and necrotic) and hyperdiploid cells could be detected after both treatments. Incubation of 2 and 3 with reduced glutathione under cell-free conditions indicated spontaneous conjugation (non-redox) reaction at pH 7.4 and pH 9.0. Analyzing the mechanism of action the total thiol content of the cells exposed to compounds 2 and 3 was determined. Compound 2 showed to reduce the total cellular thiol level both under nutrient-free and nutrient-supplemented conditions. Under the latter conditions an increase of the total thiol level of the cells exposed to 3 for 4h could be observed. The different effect of the two compounds on the cellular thiol status might contribute to the different tumor cytotoxicity of the cyclic chalcone analogues 2 and 3. Investigation of antioxidant capacity of the compounds by monitoring time course of the Fenton-reaction initiated in vitro degradation of 2-deoxyribose indicated that both compounds displayed hydroxyl radical scavenger activity. The experiments provide further details of dual--cytotoxic and cytoprotective (chemopreventive)--effects of the compounds.

Annexins↗
PubMed2005

Induction of apoptosis/necrosis in various human cell lineages by Haemophilus ducreyi cytolethal distending toxin.

We investigated the impact of highly purified Haemophilus ducreyi cytolethal distending toxin (HdCDT) on the apoptosis and necrosis of various human cells; including myeloid cells, epithelial cells, keratinocytes, and primary fibroblasts. The levels of apoptosis and necrosis induced in these cells were compared to those induced by HdCDT in human T cells and in the Jurkat T cell line. Levels of caspase-3 activity were measured, and membrane changes like phosphatidylserine (PS) translocation was evaluated after double-staining with the fluorescein isothiocyanate (FITC)-labeled annexin V and propidium iodide (PI) using flow cytometry. HdCDT induced various degrees of apoptosis and necrosis in dose- and time-dependent manners in cells of various lineages. Early and late apoptosis (annexin V-stained cells) were induced in more than 90% of T cells and monocytes after treatment with 100 ng/ml HdCDT for 24 and 48 h, respectively. The corresponding numbers for epithelial cells, keratinocytes, and fibroblasts were 26-32% after treatment with 100 ng/ml HdCDT for 48 h. HdCDT appears to eliminate effectively by inducing apoptosis those cells that are involved in immune responses. Epithelial cells, keratinocytes and fibroblasts, which are important for the healing of chancroid ulcers, are eliminated by apoptosis or necrosis after contact with HdCDT, albeit slower and to a lesser extent than T cells.

Annexin A5↗
PubMed2004

Zinc- or cadmium-pre-induced metallothionein protects human central nervous system cells and astrocytes from radiation-induced apoptosis.

We have shown the protection of human central nervous system (CNS) cultures by zinc (Zn) or cadmium (Cd)-pre-induced metallothionein (MT) synthesis from radiation-induced cytotoxicity (lactate dehydrogenase (LDH) release and neuronal dendritic injury). The present study is to further define the types of cell death induced by different dose levels of radiation and investigate the effect of MT induction (by Zn or Cd) on radiation-induced apoptosis in primary human CNS and astrocyte cultures. Apoptosis was detected by fragmented DNA electrophoresis, TUNEL technique, and propidium iodide staining. Expression of MT protein was examined by immunofluorescent staining. Results showed that exposure of primary human CNS cultures to 15 and 30 Gy gamma-radiation predominantly induced apoptotic cell death, while exposure to 60 Gy gamma-radiation predominantly induced necrotic cell death. Normal primary human CNS cultures showed weak MT staining, while primary human CNS cultures exposed to Zn or Cd showed intense MT staining. The induced apoptotic cell death by exposure to 30 Gy gamma-radiation increased to a maximum level at 12 and 24 h, and was reduced significantly by Zn or Cd pre-induced MT. Using primary human astrocytes, the induction of MT protein by Zn or Cd was further confirmed. The enhanced MT expression also afforded a significant protection from 30 Gy gamma-ray-induced apoptosis in the primary human astrocytes. These results suggest that MT protected human CNS cells from apoptosis following ionizing radiation, probably through its antioxidant property.

Apoptosis↗
PubMed2005

Evaluation of two commercial disinfectants on the viability and infectivity of Cryptosporidium parvum oocysts.

Cryptosporidiosis is mainly a problem in neonatal ruminants. Not only do Cryptosporidium spp. spread ubiquitously in our environment, but the protozoa are highly resistant to harsh environmental conditions and disinfectants, and a control measure is urgently required. This study investigated the potential biocidal activity on Cryptosporidium parvum oocysts of two commercial disinfectants developed originally to be used in farms and food-processing industries. The products, containing formaldehyde and hydrogen peroxide respectively, both had some anticryptosporidial effects. The viability and infectivity of purified C. parvum oocysts exposed to both disinfectants at different concentrations and exposure times were evaluated by inclusion or exclusion of vital dye (propidium iodide), use of an excystation technique and infection of suckling mice. Viability assays showed a decrease in oocyst viability associated with an increase in exposure time for each of the concentrations used. The intensity of infection in neonatal mice was significantly lower (P<0.05) than in the control litters.

Animals↗