In vitro correlation of ultrastructural morphology and creatine phosphokinase release in L6 skeletal muscle cells after exposure to parenteral antibiotics

Summary Morphologic changes in a rat skeletal muscle cell line (L6) exposed for 1 h to the parenteral antibiotics amphotericin B (AMP), tetracycline-HCl (TET), erythromycin lactobionate (ERY), and cephaloridine (CEP) were characterized by transmission and scanning electron microscopy and compared to cellular release of creatine phosphokinase (CRK). AMP (0.05, 0.1, 0.5 mg/ml) caused a concentration-related swelling of nuclei, endoplasmic reticulum, and mitochondria. Loss of membrane integrity associated with AMP exposure was evident at the middle concentration and extensive at the high concentration, which correlated well with the 43 and 90% depletion of CPK from the muscle cells, respectively. TET (0.25, 1.0, 2.5 mg/ml) caused dilation of endoplasmic reticulum and cytoplasmic blebbing at the low concentration but had no effect on the cytoplasmic membrane or CPK. Cells exposed to the high concentration of TET had extensive damage to the cytoplasmic membrane, and CPK was completely depleted. ERY (2.5, 5.0, 25 mg/ml) caused a pattern of morphologic changes and CPK depletion similar to TET. CEP (4.0, 20, 50 mg/ml) had no effect on membrane integrity or CPK; however, membranous whorls were prominent in the cytoplasm. A good correlation between CPK release and cytoplasmic membrane integrity was evident and the ability of these agents to release CPK from muscle cells in culture correlated with the known irritancy potential of these parenteral antibiotics. Furthermore, CPK depletion seems to be a reliable indicator of muscle cell damage after cytoplasmic membrane perturbation and is therefore an appropriate index of toxicity in this in vitro muscle irritation model.     

The human recombinant c-kit receptor ligand, rhSCF, induces mediator release from human cutaneous mast cells and enhances IgE-dependent mediator release from both skin mast cells and peripheral blood basophils.

The gene product of the steel locus of the mouse represents a growth factor for murine mast cells and a ligand for the c-kit proto-oncogene receptor, a member of the tyrosine kinase receptor class of oncogenes (for review, see O. N. Witte. 1990. Cell 63:5). We have studied the effect of the human recombinant c-kit receptor ligand stem cell factor (rhSCF) on the release of inflammatory mediators from human skin mast cells and peripheral blood basophils and compared its activity to that of rhIL-3, rhSCF (1 ng/ml to 1 microgram/ml) activated the release of histamine and PGD2 from mast cells isolated from human skin. Analysis by digital video microscopy indicated that purified human skin mast cells (84 +/- 5% pure) responded to rhSCF (0.1 to 1 microgram/ml) challenge with a rapid, sustained rise in intracellular Ca2+ levels that was accompanied by secretion of histamine. A brief preincubation (10 min) of mast cells with rhSCF (0.1 pg/ml to 1 ng/ml) significantly enhanced (100 +/- 35%) the release of histamine induced by anti-IgE (3 micrograms/ml), but was much less effective on IgE-mediated release of PGD2. In contrast, a short term incubation with rhSCF did not potentiate the secretion of histamine activated by substance P (5 microM). A 24-h incubation of mast cells with rhSCF did not affect the release of mediators induced by anti-IgE (3 micrograms/ml), probably due to receptor desensitization, rhSCF (1 ng/ml to 3 micrograms/ml) neither caused release of histamine or leukotriene C4 (LTC4) release from leukocytes of 14 donors, nor induced a rise in intracellular Ca2+ levels in purified (greater than 70%) basophils. Brief preincubation (10 min) of leukocytes with rhSCF (1 ng/ml to 3 micrograms/ml) caused an enhancement (69 +/- 11%) of anti-IgE-induced release of histamine that was significant at concentrations as low as 3 ng/ml (p less than 0.05), whereas it appeared less effective in potentiating IgE-mediated LTC4 release. In contrast, a prolonged incubation (24 h) with rhSCF (0.1 pg/ml to 100 ng/ml) did not enhance the release of histamine or LTC4 induced by anti-IgE (0.1 microgram/ml), whereas rhIL-3 (3 ng/ml) significantly potentiated the release of both mediators.(ABSTRACT TRUNCATED AT 400 WORDS)     

白粉菌侵染对小麦叶片显微及超微结构的影响白粉菌侵染对小麦叶片显微及超微结构的影响

通过半薄及超薄切片,比较了正常和受白粉菌感染的小麦叶片细胞的显微及超微结构的差异。观察结果发现(1)受感染小麦叶肉细胞的细胞壁上局部沉积大量团状电子致密颗粒;(2)叶绿体形状由原来的椭圆形转变成圆形,叶绿体膜破裂,类囊体膨大,基粒片层排列疏松,同时,叶绿体内嗜锇性颗粒数量增加;(3)线粒体膜解体,内含物分散到了细胞质中     

Gene and protein expressions in human cord blood cells after exposure to acrylonitrile

Acrylonitrile is a very high volume industrial chemical used primarily in the manufacture of plastics and rubber, which displays a pronounced acute toxicity and may be carcinogenic. The damage to the hematopoietic function by acrylonitrile may result from interference with cytokine production and cytokine receptor binding. Our present data show that acrylonitrile modulates the expression of some genes implicated in cell differentiation, cell-cycle progression, and clonogenic potential of human cord blood cells. A macroarray hybridization analysis showed that expression of the CXCR4, MCP-1, and MRP8 genes was modified by acrylonitrile exposure. Moreover, the acrylonitrile cell target seems to be the myeloid compartment, as assessed by a CFU-GM assay. In particular, the downregulation of CXCR4, MCP1, and MRP8 can be responsible for the observed reduction of cell proliferation and clonogenic capability of CFU-GM precursors. A Western blot assay showed an acrylonitrile-dependent induction of Bax, while Bcl-2 expression changed only after 48 h of chemical exposure. Bax was overexpressed in respect to Bcl-2, and this fact can be responsible for the induction in cell death after 24 h of treatment. C-fos and c-jun were also downregulated after 24 h and 6 h of treatment, respectively.     

Human sperm aneuploidy after exposure to pesticides

This study examined the effect of paternal environmental exposure to pesticides on the frequency of aneuploidy in human sperm. To determine if the chromosome number in germ cells was altered by paternal exposure, multicolor fluorescence in situ hybridization (FISH) analysis was utilized to measure aneuploidy frequencies in the sperm of 40 men (20 exposed, 20 controls). Samples were coded for "blind analysis" to eliminate scorer bias. Aneuploidy and diploidy frequencies were assessed for chromosomes 13, 21, X, and Y. A minimum of 10,000 sperm was scored per donor per chromosome probe with a total of 809,935 sperm scored. Hybridization efficiency was 99%. There were no significant differences in aneuploidy or diploidy frequencies between exposed and control groups, suggesting that the pesticides did not increase the risk of numerical chromosomal abnormalities in these men.     

Successive contrast cannot explain suppression release after repetitious exposure to one of the components of a taste mixture

Repetitious exposure to one of the components of a NcCl-sucrosemixture causes the other component to recover from suppression.This so-called‘suppression release’ might be explainedby assuming that the subjects habituate to the repeated component,which is thereby disabled as a suppressor in the mixture. Twoexperiments are reported that test successive contrast as analternative explanation of this type of suppression release.This alternative was investigated by substituting an unmixedtest stimulus of about equal subjective sweetness (or saltiness)for the mixture. The results indicate that successive contrastcontributes only partially to the suppression-release effect.When the contrast contribution is subtracted, a significantsuppression-release effect remains.     

Cytochemical and ultrastructural changes in aflatoxin-induced injury to rat liver cells

Young male Sprague-Dawley rats were injected intraperitoneally with a single dose of aflatoxin B1 (AFB1, 3 mg/kg). At 3, 6, 12, and 24 hours and 1, 2, and 5 weeks, the rats were killed and liver samples were taken for examination of sequential ultrastructural changes and localization of acid phosphatase (AcPase), thiamine pyrophosphatase (TPPase) and glucose-6-phosphatase (G6Pase) activity. At 3-6 hrs of AFB1 treatment, the nucleoli became compacted and the network forms of nucleolonema disappeared. Parallel arrays of rough ER encountered in normal liver cells became deranged. Smooth ER increased to form groups of SER anastomosis or vesicles near the golgi area. At 12-24 hours, disruptions of nucleoli, ER systems, and polysomes became more evident. Parallel arrays of ER membranes, forming whorls in the cytoplasm as well as in the cytoplasmic patches (CP), were G6Pase-positive, although the CP were AcPase-negative. The TPPase reaction in bile canaliculi was frequently diminished, but was present in some measure in the Golgi saccules. By 1-2 weeks, most of the injured cells had recovered gradually. The CP disappeared and parallel arrays of RER were observed again in most parenchymal cells. At 5 weeks, the appearance of the nucleoli was normal, as was that of the other organelles. We concluded that the hepatic parenchymal cells had serious lesions at 12 and 24 hours of AFB1 treatment and then recovered nonsynchronously. The response, resistance, and ability to recover from the toxicity of AFB1 varied among the parenchymal cells. The three marker enzymes persisted throughout all regimens of AFB1 treatment.     

Structural and ultrastructural changes in Mycobacterium rubrum cells exposed to penicillin

The action of penicillin taken at subbacterioscopic doses on Mycobacterium rubrum cells causes changes in the size and shape of the cells, in the structure of the cell wall, in the intracellular membrane systems and in functions associated with them, and in the structure of nucleoids whose DNA packing becomes more loose. If the antibiotic is added at bacteriostatic doses, the size and shape of the cells do not change, but peptidoglycan precursors being synthesized are not incorporated into the polymer and accumulate in the periplasm. DNA overspiralization in nucleoids is a non-specific reaction, which indicates that DNA is physiologically passive. DNA is isolated with a membrane from the cytoplasm in certain cells. It is possible that the resistance of cells against penicillin is associated with the capability of DNA to become inactive in physiological terms.     

Physiological and ultrastructural changes in Stellaria media following treatment with fluroxypyr

The physiological and ultrastructural changes induced by fluroxypyr (4-amino-3, 5-dichloro–6–fluoro–2–pyridyloxyacetic acid) are investigated in a susceptible weed species, Stellaria media. The sequence of symptoms was an initial petiole curvature followed by stem elongation and thickening prior to leaf senescence, stem and petiole necrosis and plant death. Light and electron microscopy revealed leaf tissue containing cells with disrupted and swollen chloroplasts and a disrupted tonoplast. Within the stem tissue there was extensive meristematic differentiation and adventitious root development prior to stem tissue necrosis. Increased levels of reducing sugars and amino acids in treated foliar tissue indicated reserve mobilisation during the initial stages of symptom development. Fluroxypyr induced oat coleoptile elongation with an optimum concentration at 5 × 10^-5 m compared to 10^-4 m for auxin (IAA). A S. media shoot explant system revealed similar elongation which was maximal during the first 24 h of treatment. Uptake and incorporation of ¹⁴C-leucine was stimulated by 96 h after treatment. It is concluded that fluroxypyr activity against S. media is characterised by an initial cell elongation and reserve mobilisation, followed by extensive cell proliferation, increased RNA and protein synthesis, and ultimately plant death.     

Calcium-dependence of catecholamine release from bovine adrenal medullary cells after exposure to intense electric fields

Summary By subjecting isolated adrenal medullary cells to intense electric fields of brief duration it is possible to gain access to the cell interior without impairing the ability of the cell to undergo exocytosis. After a single exposure to a field of 2 kV/cm, =200 sec, adrenal medullary cells behave as if their plasma membrane contains two pores of effective radius 2 nm. At 37°C these equivalent pores remain patent for up to 1 hr. The formation and stability of these pores is not affected by the Ca content of the bathing solution. The pores permit externally applied catecholamine and Ca-EGTA to equilibrate rapidly with the cell water.Cells rendered leaky in K glutamate medium containing 5mm Mg-ATP and EGTA to give an ionized Ca close to 10^–8 m release less than 1% of their total catecholamine. These same cells can release up to 30% of their catecholamine when exposed to 10^–5 m Ca. This Ca-dependent release is unaffected by Ca-channel blockers such as D600. Catecholamine release in response to a calcium challenge only seems to occur during the first few minutes whilst the Ca concentration is changing, and the extent of release depends on the final Ca concentration achieved. Half-maximal release occurs at about 1 m Ca, and this value is independent of the EGTA concentration used to buffer the ionized Ca. The relation between ionized Ca and catecholamine release is best fitted by a requirement for 2 Ca ions.Calcium-evoked release of catecholamine is associated with the release of dopamine--hydroxylase (DH) but not lactate dehydrogenase. The ratio DH/catecholamine released is the same as that in stimulated intact cells and perfused glands. The time course of appearance in the external medium of DH and catecholamine is identical. Transmission electron microscopy of leaky cells exposed to 10^–8 m Ca reveals no marked differences from unstimulated intact cells. The cytoplasm of leaky cells exposed to 10^–5 m Ca contains large membrane-bounded vacuoles. When secretion is caused to take place in the presence of horseradish peroxidase, this marker is found within the vacuoles.Ca-dependent release of both catecholamine and DH requires Mg-ATP. Cells equilibrated with Ca in the absence of Mg-ATP can be triggered to undergo exocytosis by the addition of Mg-ATP. In the absence of Mg, ATP alone is ineffective. Of a variety of other nucleotides tested, none is as effective as ATP. Mg-ATP affects the extent of exocytosis and not its apparent affinity for Ca.Replacement of glutamate as the major anion by chloride results in a marked reduction in Ca-dependent release of both catecholamine and DH. Chloride causes a small increase in Ca-independent release of catecholamine, a large reduction in the extent of exocytosis, and a decrease in the apparent affinity of exocytosis for Ca. Of a variety of anions examined, their order of effectiveness at supporting Ca-dependent exocytosis is glutamate^–>acetate^–>Cl^–>Br^–>SCN^–.Exocytosis is not obviously affected by replacing K by Na or sucrose or by altering the pH over the range pH 6.6 to 7.8. Raising the free Mg concentration reduces the extent of Ca-dependent exocytosis and also its apparent affinity for calcium. Calcium-dependent exocytosis in leaky cells is largely unaffected by (i) a variety of agonists and antagonists of the nicotinic receptor; (ii) agents that disrupt microtubules and microfilaments; (iii) phalloidin; (iv) vanadate; (v) inhibitors of anion permeability; (vi) protease inhibitors; and (vii) agents that dissipate the vesicle pH gradient and potential. It is partially inhibited by (i) certain antipsychotic drugs; (ii) a rise in osmotic pressure, (iii) lowering the temperature below 20°C, and (iv) N-ethyl maleimide.