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PubMedDate not supplied

Ototoxicity of the new platinum anticancer agent TRK-710 in comparison with cisplatin.

The pharmacokinetics and ototoxicity of the new platinum analogue TRK-710 (3, 9, 15 mg/kg x 3 days) were compared with those of cisplatin (1, 3, 5 mg/kg x 3). The perilymphatic concentration of TRK-710 was one seventh of that of cisplatin even 1 h after the administration. The N1 threshold of the compound action potential was elevated dose-dependently in both groups with a similar degree of hearing impairment. Morphological observation using phase contrast microscopy and scanning electron microscopy revealed the damage of the outer hair cells to almost the same degree mainly in the basal and second turns. Despite its usefulness against cisplatin-resistant tumors and a lesser degree of nephrotoxicity and myelosuppression, TRK-710 should be clinically used with caution similar to cisplatin.

Action Potentials↗

PubMed2004

Platelet-leukocyte aggregation induced by PAR agonists: regulation by nitric oxide and matrix metalloproteinases.

Platelet-leukocyte aggregation (PLA) links haemostasis to inflammation. The role of nitric oxide (NO) and matrix metalloproteinases (MMP-1, -2, -3, -9) in PLA regulation was studied. Homologous human platelet-leukocyte suspensions were stimulated with thrombin (0.1-3 nM) and other proteinase activated receptor-activating peptides (PAR-AP), including PAR1AP (0.5-10 microM), PAR4AP (10-70 microM), and thrombin receptor-activating peptide (1-35 microM). PLA was studied using light aggregometry with simultaneous measurement of oxygen-derived free radicals, dual colour flow cytometry, and phase-contrast microscopy. The release of NO was measured using a porphyrinic nanosensor, while MMPs were investigated by Western blot, substrate degradation assays, immunofluorescence microscopy, and flow cytometry. The levels of P-selectin and microparticles (MP) in PLA were measured by flow cytometry. PLA was also characterized using pharmacological agents: S-nitroso-glutathione (GSNO, 0.01-10 microM), 1H-Oxadiazole quinoxalin-1-one (ODQ, 1 microM), N(G)-L-nitro-L-arginine methyl ester (L-NAME, 100 microM) and compounds that modulate the actions of MMPs such as phenanthroline (100 microM), monoclonal anti-MMP antibodies, and purified MMPs. PAR agonists concentration-dependently induced PLA, an effect associated with the release of microparticles (MP) and the translocation of P-selectin to the platelet surface. NO and radicals were also released during PLA. Inhibition of NO bioactivity by the concomitant release of free radicals or by the treatment with L-NAME or ODQ stimulated PLA, while pharmacological administration of GSNO decreased PLA. PAR agonist-induced PLA resulted in the liberation of MMP-1, -2, -3, and -9. During PLA, MMPs were present on the cell surface, as shown by flow cytometry and immunofluorescence. PLA led to the activation of latent MMPs to active MMPs, as shown by Western blot and substrate degradation assays. Inhibition of MMPs actions by phenanthroline and by the antibodies attenuated PLA. In contrast, purified active, but not latent, MMPs amplified thrombin-induced PLA. It is concluded that NO and MMP-1, -2, -3, and -9 play an important role in regulation of PAR agonist-induced PLA.

Blotting, Western↗

PubMed1986

Development of cultured cardiac cells and difference of their structures in vivo.

Some interesting features in the morphological study of rat cardiac cells were obtained by use of the light and phase-contrast microscopy. The histological approach and comparison of the ultrastructures of the cultured cardiac cells with uncultured cells were included in the studies as a control. Four main points are discussed: the process of single beating cells that eventually constitutes the cell cluster and network, the function and histological content of the network, the mechanisms of synchronized beating cells, the substance of the cell surface coat and intercellular connection.

Animals↗

PubMed2006

Mechanical energy from intraocular instruments cause emulsification of silicone oil.

AIM: The complications of intraocular silicone oil include emulsification, which may lead to vision-threatening disorders such as keratopathy, secondary glaucoma and retinopathy. The authors study the effect of mechanical energy from surgical instruments on the emulsification of silicone oil. METHODS: Three different handpieces (phacofragmentation, phacoemulsification, high-speed vitrectomy) were placed at the interface of balanced salt solution and silicone oil (1000 and 5000 cst, 200 fluid; Dow Corning). The phacofragmentation handpiece was evaluated over different ultrasound powers and duration. Emulsification was quantified with phase-contrast microscopy and manual counting of digital photographs by a masked examiner. In addition, phacoemulsification was performed in the anterior chamber of a human ex vivo eye with full-fill silicone oil. RESULTS: Emulsification increases with higher phacofragmentation power and duration and is greater for 1000 cst silicone oil. Emulsification of silicone oil occurs with phacoemulsification and high-speed vitrectomy handpieces. CONCLUSIONS: The energy from surgical handpieces is sufficient to result in silicone oil emulsification.

Emulsions↗

PubMed2000

Cell fate decisions in a human retinal precursor cell line: basic fibroblast growth factor- and transforming growth factor-alpha-mediated differentiation.

The purpose of this study was to determine if immortalized human retinal precursor cells could serve as a model to investigate cues that modulate cell fate and differentiation. We investigated the effects of a variety of growth factors broadly but specifically tested the effects of basic fibroblast growth factor (bFGF) and transforming growth factor (TGF)a in retinal cell differentiation and commitment. To determine the role of exogenously added growth factors in a human retinal precursor cell line (KGLDMSM), established from a first-trimester retina, cells were adapted to grow in a defined medium and exposed to a variety of trophic factors (epidermal growth factor [EGF], neuron growth factor [NGF], TGFalpha, TGFbeta, acidic FGF, and bFGF). Dose-response curves were developed to arrive at optimal concentrations. The neurotrophic potential of growth factors was determined by 3H-thymidine incorporation and bromodeoxyuridine (BrdU) labeling. The identity of the emerging neuronal phenotypes were determined by phase-contrast microscopy, immunolabeling for the neuron-specific antigens neurofilament protein (NF) and neuron-specific enolases (NSE), and photoreceptor-specific antigens (Rho1D4, 7G6) using immunocytochemistry and Western blot analysis. To identify some of the early response genes (c-fos, c-myc) expressed in response to growth factors, Northern blot analysis was performed. Almost all of the factors tested increased the total number of cells with a neuronal phenotype. Potency of growth factors to generate neurons was TGFalpha > bFGF > EGF > NGF. Both TGFalpha and bFGF, alone or in combination, increased the total number of neurons. Most of the neurons generated were photoreceptors, as depicted by the polarized phenotype, expression of photoreceptor-specific antigens, and processes resembling rudimentary outer segments. The increase in photoreceptor-like neurons is possibly attributable to an increase in numbers rather than greater survival. Additionally, the majority of the photoreceptors generated labeled with BrdU and for photoreceptor-specific antigens, suggesting that an inductive effect of bFGF and TGFalpha could occur in the cell cycle or shortly thereafter. Both bFGF and TGFalpha induced the expression of the early response gene c-fos while not altering the expression of c-actin or c-myc. The emergence of a photoreceptor phenotype was confirmed by both immunocytochemistry and Western blot analysis. The immortalized retinal precursor cell line could prove valuable in determining the role of exogenously added growth factors in retinal development and differentiation. Both bFGF and TGFalpha enhance the photoreceptor phenotype in medium-density cultures under conditions of defined medium. The same was confirmed by phase-contrast microscopy, immunocytochemistry, and Western blot analysis. Furthermore, cell fate determination in cultured precursor cells could occur during the late part of the cell cycle or shortly after completion of cell division. The effects of TGFalpha and bFGF seem to be slightly additive. The cell line will be extremely valuable in studying mechanisms of cell commitment and generation of retinal cell types, which could be tested for their potential for transplantation.

Cell Differentiation↗

PubMed1979

Effects of lithium on morphological characteristics of dissociated brain cells in culture.

Lithium chloride was added in 5 and 10 mM concentrations for different periods of exposition time to dissociated cultures obtained from chicken embryonic brain. When supplementing lithium at day 1 in vitro for five days, a dose-dependent decrease in total protein was observed in the cultures as compared to the sodium-treated controls. Profound reduction was revealed in the length of neuronal processes and in the number of neuronal cell bodies by phase contrast microscopy and by morphometric means. After exposition of lithium in 10 mM concentration for 48 h, beside a slight decrease in number of perikaryons, a marked reduction in process length of neural elements was observed in the 6-day-old tissue cultures. Ultrastructurally, swollen and degenerating nerve processes have been found after lithium treatment suggesting a particular sensitivity of these structures to lithium ion.

Animals↗

PubMed1980

Evidence suggesting a role for cyclic nucleotides in acrosome reactions of hamster sperm in vitro.

There have been conflicting reports concerning the involvement of cyclic nucleotides in sperm capacitation. We have examined the effects of micromolar concentrations of dibutyryl cyclic AMP (Bt2cAMP) and of the phosphodiesterase inhibitors SQ20009 and ICI63,197 on hamster sperm incubated under in vitro capacitating conditions. Washed hamster sperm were incubated in a capacitation media containing bovine serum albumin, and a protein-free "motility-factor" from bovine adrenal cortex. Incubation for 3.5 hours was followed by addition of one of the compounds (0.1-10 microM) or control buffer. At the time of addition and after 30-120 minutes further incubation, sperm were examined by phase contrast microscopy. The final motility was similar to the initial motility (50-70%) and the same in incubation of controls or experimental compounds. Bt2cAMP, SQ20009, and ICI63,197 at these concentrations stimulated acrosome reactions to a statistically significant extent (P less than 0.005) compared to controls. Activation was stimulated to a varying degree by all three experimental compounds. These results suggest a role for cyclic nucleotides in capacitation and the acrosome reaction of hamster sperm.

Acrosome↗

PubMed1981

Organotypic monolayer cultures of nervous tissue.

Cultivation of nervous tissue by means of the roller-tube technique yields thin organotypic cultures. Explants or slices prepared from 1- to 20-day-old rats are embedded in a plasma clot on flying coverslips and cultivated for weeks in roller-tubes. Due to the flattening of the tissue, individual nerve cells are often arranged in monolayer thickness and can, therefore, be viewed with phase-contrast microscopy. This technique is utilized to culture and co-culture nervous tissue derived from various brain regions. The degree of organotypic organization depends on the age of the animals used for culturing. Stable intracellular recordings arae obtained from nerve cells which are impaled under visual control. In view of the accessibility of individual living cells, this approach seems to be particularly well-suited for physiological and pharmacological studies on morphologically identified nerve cells.

Animals↗

PubMed1965

PREPARATION AND USE OF FIXATIVE SLIDES FOR THE DIAGNOSIS OF TRICHOMONAS VAGINALIS.

Clean microscopic slides are dipped in a solution of mercuric chloride and sucrose in 30% alcohol and allowed to dry. A thin transparent film of cytological fixative is thus formed on both sides of the slide. Fixative slides, prepared in this manner, can be stored indefinitely provided that they are not exposed to a warm, humid atmosphere. When a drop of fluid containing bacteria, protozoa, or mammalian cells is spread over the surface of such a slide the formed elements are immediately fixed, the finer cytological details being well preserved. Staining and examination are carried out when the specimen arrives at the laboratory. This method is particularly suitable for a diagnostic laboratory service in which most of the specimens arrive by mail. The method has been successfully employed in the diagnosis of Trichomonas vaginalis infection of the female genital tract. This parasite was found by phase-contrast microscopy in 47% of 1,000 women examined at a police court clinic. Fixative slide preparations were also obtained from these women at the time of their medical examination. Of these specimens, 38.6% showed the presence of Tr. vaginalis, and 10% were also found to be infected with N. gonorrhoeae. The fixative slide technique may also prove of value in such widely separated fields as exfoliative cytology and the diagnosis of primary syphilis.

Animals↗

PubMed1989

Identification of Tritrichomonas foetus in sections of bovine placental tissue with monoclonal antibodies.

Sections of bovine placenta from cases of bovine trichomoniasis were examined for the presence of Tritrichomonas foetus by standard histological methods using phase-contrast microscopy and by indirect immunofluorescence assay (IFA) employing monoclonal antibodies (mAbs) specific for T. foetus. Parasites were identified readily in deparaffinized tissue up to 4 yr old by IFA with 2 mAbs previously shown to bind to the surface of living T. foetus. These results indicated that the IFA provided a rapid and specific method of identifying T. foetus in tissue sections as compared to standard histological methods.

Abortion, Veterinary↗

PubMed1989

[Rapid methods of detection of bacteria in the bile].

Rapid methods are described for assessing the bacterial contamination, using direct phase-contrast microscopy, triphenyltetrazolium chloride (TTC) test, and a standard loop 3 mm in diameter, or by the detection of the biliary 'active' leukocytes and the microflora sensitivity to antibacterial drugs with the use of the TTC test and turbidimetric technique. These methods permit getting an answer in 4-24 hrs, are simple, and do not require special bacteriologic equipment, this recommending them for routine clinical studies.

Bacteria↗

PubMed1986

Techniques for examining Pneumocystis carinii in fresh specimens.

Pneumocystis carinii was examined in fresh preparations of infected rat lung homogenates and tissue culture supernatants by a variety of light microscope techniques, vital dyes, and histologic stains. Phase-contrast microscopy, Nomarski interference-contrast microscopy, and bright-field microscopy with oblique illumination provided excellent views of P. carinii. Erythrosin B, and to a lesser extent trypan blue, were helpful in assessing organism viability. The use of Triton X-100-Giemsa stain permitted differentiation of the developmental stages in the P. carinii life cycle. The techniques developed here are easily adaptable to the microbiology laboratory and thus should have important clinical and research applications.

Animals↗

PubMed1996

Isolation and in vitro propagation of human choroidal fibroblasts. Morphology and characterization of the cultures.

Subretinal ingrowth of choroidal fibroblasts is of importance in several clinical settings such as age-related macular degeneration or after traumas such as laser treatment and infection. In the present investigation we describe a method for isolation and propagation of human choroidal fibroblasts in vitro. Outgrowth from the outseeded choroid was studied by means of phase contrast microscopy, scanning electron microscopy and by immunohistochemical methods. The secondary cultures were found to contain almost only fibroblasts (more than 95% of the cells), some contaminating retinal pigment cells were also found. Choroidal fibroblasts contribute to the fibrovascular subretinal scarring process, and our cell culturing system seems to be a suitable tool for studying factors regulating the behavior of these cells in vitro.

Aged↗

PubMed1975

Stoichiometry of compounds bound to human erythrocytes in relation to morphology.

Most work on human erythrocyte interaction with drugs and other compounds has been reported on the basis of total concentrations. Total concentrations alone do not reveal numbers of molecules bound per cell, v. This paper emphasizes determination of v and of binding isotherms, in conjunction with changes in cell morphologies and in hypotonic shock behavior as v is varied. Four drugs and five other compounds were studied, with fresh erythrocytes. The principal findings are: (1) the intact erythrocyte engages in two kinds of binding mechanisms, statistical binding and cooperative binding, depending on the compound. In the case of a detergent, dodecylbenzene sulfonate, the binding is nearly quantitative. (2) The compounds often induce considerable protection against hypotonic hemolysis. However, the binding levels at which maximum protection occurs are rather close to the levels, vL, that occur upon complete conversion to the first distorted morphology. Therefore, the maximally protected erythrocyte may be a distorted erythrocyte. (3) The value n is the apparent total number of sites from Scatchard plotting for compounds which bind in a statistical manner. Levels vp and vw characterize maxima in cooperative binding behavior, also from Scatchard plotting of the data. Despite the wide diversity of over-all levels at which compounds exert their effects, the critical binding levels of and numbers of sites fall into a narrow range:n, vL, vP, and vw are all between 1 and 8 times 10-7 molecules or sites per cell. Most of our data, and that from some other laboratories, indicate that about 2 plus and minus 1 times 10-7 sites per erythrocyte are available for compound binding by the intact cell. Beyond that level, the cell in suspension almost always will be forced into the first obvious morphology change, as seen by phase contrast microscopy. (4) Once stoichiometries are established, the total binding capacity of erythrocytes for such compounds, in blood, can be estimated. An intruding organic molecule would encounter about 6 times as many plasma albumin sites as erythrocyte sites, if the plasma albumin sites were free. However, because albumin in vivo usually forms a complex with one to two fatty acids, the erythrocyte itself is rather likely to act as a transport particle for such compounds.

Binding Sites↗

PubMed1975

Antibiotic-resistant mutants of Bacillus subtilis conditional for sporulation.

Among spontaneously occurring antibiotic-resistant mutants of Bacillus subtilis 168 we have identified a sub-class that is conditionally sporulative. Mutants in this sub-class are resistant to antibiotic during vegetative growth but are sensitive during sporulation. Mutants conditionally-resistant to erythromycin, kanamycin, spectinomycin, and streptomycin have been isolated and characterized by phase contrast microscopy and with respect to their ability to synthesize heat-resistant endospores or the sporulation-associated enzyme alkaline phosphatase. The results suggest that several entirely different genetic lesions may result in this single phenotype. This group includes mutants whose properties suggest that both th 30S and 50S ribosomal subunits may be altered concomitant with early spore specific metabolism. The blockage imposed by antibiotic may be at or near Stage 2 of sporulation.

Anti-Bacterial Agents↗

PubMed1977

Reticulocyte motility and form: studies on maturation and classification.

A new two-stage living cell cytological classification is presented for reticulocytes in the rat and shown on an overall population basis to be the equivalent of the classical new methylene blue staining characterization. Class 1 reticulocytes (R1) are motile and multilobular. They comprise about 25% of normal bone marrow reticulocytes, but none of the reticulocytes in normal blood. Class 2 reticulocytes (R2) are asymmetrically "deep dished" in appearance and contain visible refractile granules, viewed in phase-contrast microscopy. In animals "stimulated" by bleeding, the percentage of motile marrow reticulocytes is found to be a linear function of hematocrit over the range of about 25%-70% total reticulocytes. In the blood, the associated maximum rise of class R1 is only to about 8%. These and related cellular properties are discussed with respect to the maturation and release of normal reticulocytes. Applicability of this work to studies on humans and to pathology is also indicated.

Animals↗

PubMed2001

Protein kinase C alpha and gamma in N/N 1003A rabbit lens epithelial cell differentiation.

PURPOSE: To determine if Protein Kinase C (PKC) plays a role in the initiation of lens epithelial cell differentiation into a lens fiber cell. METHODS: PKCalpha or PKCgamma was overexpressed in N/N 1003A lens epithelial cells for up to 7 days. Phase contrast microscopy was used to observe morphological changes associated with PKCalpha or PKCgamma overexpression. Cell cycle changes in cells overexpressing PKCalpha or PKCgamma were measured using acridine orange staining and flow cytometry. Crystallin levels in cells overexpressing PKCalpha or PKCgamma were measured using Western blots and RT-PCR. RESULTS: Significant differences in cell cycling were observed between untransfected cells and those overexpressing PKCalpha or PKCgamma. Overexpression of PKCalpha and PKCgamma caused the cells to lose their epithelial-like appearance and elongate. alphaB-crystallin expression was detected in all the samples while alphaA-crystallin was detected only in cells after 7 days of PKCalpha or PKCgamma overexpression. CONCLUSIONS: The observations that alphaA-crystallin is only found in N/N 1003A cells overexpressing PKCalpha or PKCgamma for 7 days along with the finding that a block in the G0/G1 phase of the cell cycle and the consequent morphological changes are observed, indicate that PKCalpha and PKCgamma may have a role in the initiation of differentiation in lens epithelial cells.

Acridine Orange↗

PubMed1984

[Cytomorphology of the basic histological forms of thymus tumors (cytohistological parallels)].

Cytomorphology of various histological types of thymic tumours was studied on transthoracic punctures and scrapes from the operatively removed thymomas. Cytological material was stained by Pappenheim with hematoxylin and eosin, by modified Papanicolau' method as well as by means of luminescent and phase-contrast microscopy. Histological slides were stained with hematoxylin and eosin. Cytological diagnostics of thymomas is based on the detection of the two main types of cells--lymphoid and epithelial. Hassals' bodies are detectable very rarely in the material of lymphoid thymomas punctures. The cytological features characteristic of lymphoid, lymphoepithelial, epithelial, and spindle-cell thymoma are described. The identity of cell types in both cytological smears and histological slides is stated, taking into account their changes in these two research methods.

Biopsy, Needle↗