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Biomedical subjects

Olaf Strauss

Publications and source records attributed to Olaf Strauss.

14 recordsLinked to original sources
PubMed2007

Lack of antioxidative properties of vitamin C and pyruvate in cultured retinal pigment epithelial cells.

BACKGROUND: Oxidative damage to the retinal pigment epithelium might be involved in the pathogenesis of age related macular degeneration. Thus antioxidative protection represents a rationale for a causative therapy or prophylaxis. The aim of the present study is to evaluate antioxidative properties of vitamin C and pyruvate at retinal pigment epithelial (RPE) cells exposed to oxidative stress. METHODS: The ability of vitamin C and pyruvate to quench hydroxyl radicals was tested using the di-hydro-rhodamine (DHR) assay. Cells of the human RPE cell line ARPE-19 were exposed for 8 min to hydroxyl radicals generated by the Fenton reaction from 2.25 mM H2O2 and 30 microM Fe3+ -nitrilo-tri-acetate. This was done in the absence and presence of 0.3-3.0 mM pyruvate and vitamin C, respectively. Cell survival was analysed by vitality staining (life-dead-assay) and expressed as cell survival ratio. A survival ratio <1.0 indicates cell loss. RESULTS: At concentrations from 0.1 to 1.0 mM vitamin C and pyruvate quench hydroxyl radicals in the DHR assay in absence of living matter. In the presence of 0.1- 0.3 mM vitamin C and pyruvate, ARPE-19 showed a reduced survival ratio (0.87 +/- 0.01 to 0.89 +/- 0.02 after 6 h) which was not the case at the higher concentrations between 1 and 3 mM. The exposure of ARPE-19 cells to hydroxyl radicals reduced the survival ratio to 0.92 +/- 0.02. At concentrations at which vitamin C and pyruvate exert toxic effects, a potentiation of radical induced cell death can be observed (survival ratio 0.79 +/- 0.02 and 0.82 +/- 0.03, respectively). Higher concentrations of vitamin C or pyruvate had no explicit protective effect to the hydroxyl radical induced damage. DISCUSSION: Although vitamin C and pyruvate are potent hydroxyl radical quenchers in vitro they failed to protect cultured ARPE-19 cells from oxidative stress induced cell death. In contrast, when applying the scavengers at low concentrations a potentiation of cell damage was observed.

Antioxidants↗
PubMed2006

Insulin-like growth factor-1 increases intracellular calcium concentration in human primary neuroendocrine pancreatic tumor cells and a pancreatic neuroendocrine tumor cell line (BON-1) via R-type Ca2+ channels and regulates chromogranin a secretion in BON-1 cells.

Insulin-like growth factor 1 (IGF-1) is a potent mitogenic and secretory factor that acts on voltage operated Ca(2+) channels (VOCCs). VOCCs are categorized into L-type channels (Ca(V)1.1-1.4), P/Q-type channels (Ca(V)2.1), N-type channels (Ca(V)2.2), R-type channels (Ca(V)2.3), and T-type channels (Ca(V)3.1-3.3). Aside from regulating membrane excitability, VOCCs influence chromogranin A (CgA) secretion in neuroendocrine tumor (NET) cells. It is not known, whether VOCCs play a role in the IGF-1-dependent regulation of CgA secretion in NET cells. We therefore studied the effects of IGF-1 on individual VOCC subtypes and characterized their role in mediating IGF-1-dependent regulation of CgA secretion in NET cells. Using specific modulators of VOCC subtypes, we identified the functional expression of L-, N-, P/Q- and R-type channels in primary as well as permanent models of NET. The IGF-1-induced intracellular Ca(2+) increase in NET cells was mainly due to the activation of R-type channel activity. The effects on intracellular calcium, observed in whole-cell patch-clamp recordings and fluorescence imaging, were partially blocked by the specific R-type channel blocker SNX-482 and antisense oligonucleotides against the alpha(1) subunit of this channel. IGF-1 potently induced CgA secretion. The effect of IGF-1 was reduced by both, inhibition of R-type channel activity and a reduction of R-type channel expression using antisense oligonucleotides. Since R-type channels exist in NET cells and couple to both, IGF-1 receptor signaling as well as CgA secretion, pharmacological interference with R-type channels may represent a new therapeutic option by blocking Ca(2+) signaling thereby abrogating IGF-1-dependent hypersecretion in NET disease.

Biomarkers, Tumor↗
PubMed2006

The light peak of the electroretinogram is dependent on voltage-gated calcium channels and antagonized by bestrophin (best-1).

Mutations in VMD2, encoding bestrophin (best-1), cause Best vitelliform macular dystrophy (BMD), adult-onset vitelliform macular dystrophy (AVMD), and autosomal dominant vitreoretinochoroidopathy (ADVIRC). BMD is distinguished from AVMD by a diminished electrooculogram light peak (LP) in the absence of changes in the flash electroretinogram. Although the LP is thought to be generated by best-1, we find enhanced LP luminance responsiveness with normal amplitude in Vmd2-/- mice and no differences in cellular Cl- currents in comparison to Vmd2+/+ littermates. The putative Ca2+ sensitivity of best-1, and our recent observation that best-1 alters the kinetics of voltage-dependent Ca2+ channels (VDCC), led us to examine the role of VDCCs in the LP. Nimodipine diminished the LP, leading us to survey VDCC beta-subunit mutant mice. Lethargic mice, which harbor a loss of function mutation in the beta4 subunit of VDCCs, exhibited a significant shift in LP luminance response, establishing a role for Ca2+ in LP generation. When stimulated with ATP, which increases [Ca++]I, retinal pigment epithelial cells derived from Vmd2-/- mice exhibited a fivefold greater response than Vmd2+/+ littermates, indicating that best-1 can suppress the rise in [Ca2+]I associated with the LP. We conclude that VDCCs regulated by a beta4 subunit are required to generate the LP and that best-1 antagonizes the LP luminance response potentially via its ability to modulate VDCC function. Furthermore, we suggest that the loss of vision associated with BMD is not caused by the same pathologic process as the diminished LP, but rather is caused by as yet unidentified effects of best-1 on other cellular processes.

Adenosine Triphosphate↗
PubMed2005

Voltage-dependent ion channels in the mouse RPE: comparison with Norrie disease mice.

We studied electrophysiological properties of cultured retinal pigment epithelial (RPE) cells from mouse and a mouse model for Norrie disease. Wild-type RPE cells revealed the expression of ion channels known from other species: delayed-rectifier K(+) channels composed of Kv1.3 subunits, inward rectifier K(+) channels, Ca(V)1.3 L-type Ca(2+) channels and outwardly rectifying Cl(-) channels. Expression pattern and the ion channel characteristics current density, blocker sensitivity, kinetics and voltage-dependence were compared in cells from wild-type and Norrie mice. Although no significant differences were observed, our study provides a base for future studies on ion channel function and dysfunction in transgenic mouse models.

Animals↗
PubMed2005

The fibroblast growth factor receptors, FGFR-1 and FGFR-2, mediate two independent signalling pathways in human retinal pigment epithelial cells.

To examine the effects and potential implications for the expression of the two basic fibroblast growth factor (bFGF) receptors, FGFR-1 and FGFR-2, in retinal pigment epithelial (RPE) cells, bFGF-dependent changes in gene expression and RPE cell function were studied. bFGF increased L-type Ca2+ channel activity of RPE cells, which in turn resulted in an increase of vascular endothelial growth factor A (VEGF-A) secretion from RPE cells. Also, both bFGF and direct stimulation of L-type Ca2+ channels by BayK8644 increased the expression of c-fos in RPE cells, to the same extent. bFGF-induced-c-fos expression was reduced by inhibition of FGFR-1, but not by L-type Ca2+ channel inhibition, demonstrating that stimulation of FGFR-1 results in a Ca2+ channel-independent change of gene expression. In contrast, stimulation of FGFR-2 results in a Ca2+ channel-dependent stimulation of VEGF secretion. Furthermore, immunohistological investigation of neovascular tissues obtained from patients with age-related macular degeneration (AMD) revealed FGFR-1 and FGFR-2 expression in the RPE of the diseased tissue. Our findings support the hypothesis that there are two different FGFR-1- and FGFR-2-dependent pathways that modulate the role of bFGF in induction of neovascularisation in AMD.

Calcium Channels, L-Type↗
PubMed2005

Expression of bestrophin-1, the product of the VMD2 gene, modulates voltage-dependent Ca2+ channels in retinal pigment epithelial cells.

Mutations in the VMD2 gene cause Best's disease, an inherited form of macular degeneration. The reduction in the light-peak amplitude in the patient's electro-oculogram suggests that bestrophin-1 influences the membrane conductance of the retinal pigment epithelium (RPE). Systemic application of the L-type Ca2+ channel blocker nimodipine reduced the light-peak amplitude in the rat electroretinogram but not a- and b-waves. Expression of bestrophin-1 in a RPE cell line (RPE-J) led to changes in L-type channel properties. Wild-type bestrophin-1 induced an acceleration of activation kinetics of Ba2+ currents through L-type Ca2+ channels and a shift of the voltage-dependent activation to more negative values, closer to the resting potential of RPE cells. Expression of bestrophin-1 with Best disease-causing mutations led to comparable shifts in voltage-dependent activation but different effects on activation and inactivation kinetics. Bestrophin W93C exhibited slowed activation and inactivation, and bestrophin R218C accelerated the activation and inactivation. Thus, transfection of RPE cells with bestrophin-1 distinctively changed L-type Ca2+ channel kinetics and voltage-dependence. On the basis of these data, we propose that presence of bestrophin-1 influences kinetics and voltage-dependence of voltage-dependent Ca2+ channels and that these effects might open new ways to understand the mechanisms leading to retinal degeneration in Best's disease.

Animals↗
PubMed2005

Effects of unoprostone and endothelin 1 on L-type channel currents in human trabecular meshwork cells.

BACKGROUND: The trabecular meshwork (TM) is a smooth muscle-like tissue with contractile properties and by this mechanisms involved in the regulation of aqueous humor outflow. Isopropyl unoprostone (Rescula, Novartis Ophthalmics), a synthetic docosanoid, reduces intraocular pressure in glaucoma patients and normal subjects. In isolated TM strips, unoprostone reduces TM contractility in the presence of endothelin 1 (ET-1). However, the signal transduction pathway of unoprostone still remains unclear. Since L-type channel currents are known to influence the contractility of TM, we examined the effects of unoprostone and ET-1 on L-type channel currents of TM cells. METHODS: The effects of unoprostone, ET-1 and the tyrosine kinase inhibitor herbimycin A on L-type channel currents of cultured human TM cells were investigated using the perforated patch configuration of the patch-clamp technique. RESULTS: Application of ET-1 had no effect on L-type channel currents. Unoprostone led to a dose-dependent reduction of control currents. The effect of unoprostone is independent of ET-1. After preincubation of cells with herbimycin A, unoprostone had no effect on the L-type channel current amplitude. Human TM cells preincubated with herbimycin A showed a reduced current density compared with control cells. Both substances, unoprostone and herbimycin A, increased the inactivation time constant of L-type channel currents. CONCLUSION: We conclude that unoprostone reduces the activity of L-type Ca2+ channels. This effect seems to be independent of ET-1. The signal transduction pathway seems to be mediated by tyrosine kinases.

Benzoquinones↗
PubMed2005

The retinal pigment epithelium in visual function.

Located between vessels of the choriocapillaris and light-sensitive outer segments of the photoreceptors, the retinal pigment epithelium (RPE) closely interacts with photoreceptors in the maintenance of visual function. Increasing knowledge of the multiple functions performed by the RPE improved the understanding of many diseases leading to blindness. This review summarizes the current knowledge of RPE functions and describes how failure of these functions causes loss of visual function. Mutations in genes that are expressed in the RPE can lead to photoreceptor degeneration. On the other hand, mutations in genes expressed in photoreceptors can lead to degenerations of the RPE. Thus both tissues can be regarded as a functional unit where both interacting partners depend on each other.

Animals↗
PubMed2004

Insulin-like growth factor-1 contributes to neovascularization in age-related macular degeneration.

Choroidal neovascularization (CNV) is a debilitating complication of age-related macular degeneration and a leading cause of vision loss. Along with other angiogenic factors like vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF)-1 and its receptor, IGF-1R, have been implicated in CNV. IGF-1 is produced in neurons and retinal pigment epithelium (RPE) but its targets and impact in CNV are not understood. IGF-1 immunoreactivity was abundant throughout surgically isolated human CNV tissues and RPE cells were immunopositive for IGF-1R. Cultured RPE cells obtained from CNV tissues expressed IGF-1R. IGF-1 stimulation of cultured cells from CNV tissues induced monophasic sustained rises in intracellular free Ca(2+). VEGF concentration in the medium of unstimulated RPE cell cultures from CNV tissues increased with time to a steady-state (8h) which was increased twofold by IGF-1 stimulation. Thus, in RPE cells IGF-1 stimulates the second messenger Ca(2+) and increases VEGF secretion which, in turn, induces neovascularization.

Aged↗
PubMed2003

Electrophysiological response of cultured trabecular meshwork cells to synthetic ion channels.

The response of living cells of the trabecular meshwork to synthetic ion channels is described. The THF-gramicidin hybrids THF-gram and THF-gram-TBDPS as well as a linked gA-TBDPS and gramicidin A were applied to cultured ocular trabecular meshwork cells. THF-gram application (minimal concentration, 10(-8) M; saturation, 10(-7) M) led to an additional conductance which displayed characteristics of weak Eisenman-I-selective cation channels, no cell destruction, an asymmetric change of the inward/outward currents, and higher current densities using Cs(+) as charge carrier compared to Na(+) and K(+). Linked-gA-TBDPS showed at 10(-12) M increases of the membrane conductance comparable to gA at 10(-7) M and a much faster response of the cells. Thus, THF-gramicidin hybrids form a basis for the use of synthetic ion channels in biological systems, which eventually may lead to new therapeutic approaches.

Animals↗
PubMed2002

Stimulation of L-type Ca(2+) channels by increase of intracellular InsP3 in rat retinal pigment epithelial cells.

The purpose of this study was to investigate the role of voltage-dependent L-type Ca(2+)channels in intracellular Ca(2+)signaling of the retinal pigment epithelium (RPE). Patch-clamp techniques in conjunction with measurements of the intracellular free Ca(2+)using the Ca(2+)-sensitive fluorescence dye fura-2 were performed using cultured rat RPE cells. Intracellular application of inositol-1,4,5-trisphosphate (InsP3; 10 microM) via the patch-pipette during the whole-cell configuration led to an increase in the intracellular free Ca(2+)([Ca(2+)](i)). This effect could be reduced by the L-type Ca(2+)channel blocker nifedipine (2 microM). At the moment of the maximal rise in [Ca(2+)](i)L-type currents displayed an increase in the current density and shifts in the activation curve and of the steady-state inactivation. Comparable changes of L-type channel activity could be observed by induction of capacitative Ca(2+)entry, a maneuver to release Ca(2+)from intracellular Ca(2+)stores independently from InsP3. The increase in L-type Ca(2+)channel activity and [Ca(2+)](i)by intracellular application of InsP3 or induction of capacitative Ca(2+)entry could be inhibited by blocking tyrosine kinase activity using genistein (5 microM) or tyrphostin 51 (10 microM). It is concluded that L-type Ca(2+)channels are involved in the Ca(2+)/InsP3 second messenger system by generating an influx of extracellular Ca(2+)into the cell. This is enabled by depletion of cytosolic Ca(2+)stores and tyrosine kinase-dependent activation of L-type channels.

Animals↗
PubMed2002

Effects of protein kinase C on delayed rectifier K+ channel regulation by tyrosine kinase in rat retinal pigment epithelial cells.

PURPOSE: Investigation of the regulation of delayed rectifier potassium channels by protein kinases in retinal pigment epithelial (RPE) cells. METHODS: Membrane currents of cultured RPE cells were measured in either the perforated-patch or whole-cell configuration of the patch-clamp technique. RESULTS: Different potassium channel blockers from animal venoms (charybdotoxin, hanatoxin, agitoxin, tityustoxin) were used to identify the delayed rectifying outward current in RPE cells as current through potassium channels, mainly composed of Kv1.3 subunits. Extracellular application of a tyrosine kinase blocker, such as genistein (100 microM), lavendustin A (10 microM), or herbimycin A (4 microM), almost completely abolished the delayed rectifier currents. Intracellular application of the tyrosine kinase pp60(c-src) (30 U/mL) through a patch pipette led to an increase of the delayed rectifier current consistent with an activation of the delayed rectifier channels by src subtype tyrosine kinase. Inhibition (with 1 microM chelerythrine) or activation (with 1 microM phorbol ester PMA) of protein kinase C (PKC) did not change the activity of delayed rectifier channels. Inhibition of PKC also did not change the effect of tyrosine kinase inhibition of delayed rectifier currents. However, in cells with stimulated PKC, inhibition of tyrosine kinase by genistein led to an increase of delayed rectifier currents. Intracellular application of pp60(c-src) (30 U/mL) in cells with stimulated PKC led to inhibition of delayed rectifier currents. CONCLUSIONS. Tyrosine kinase, of the src family, can activate or inhibit delayed rectifier channels composed of Kv1.3 subunits in RPE cells. The activity of PKC determines whether tyrosine kinase activates or inhibits these K+ channels.

Animals↗
PubMed2002

Ca2+-channels in the RPE.

The retinal pigment epithelium closely interacts with photoreceptors and helps to maintain the activity of photoreceptors. Investigations using patch-clamp techniques on cultured or freshly isolated retinal pigment epithelial cells from various species demonstrated the expression of voltage-dependent Ca2+ channels with characteristics of L-type channels. Since retinal pigment epithelial cells rarely display changes of the membrane potential which lead to the activation of these Ca2+ channels, their function seemed to be unclear. Recent findings shed light onto the possible role of these Ca2+ channels. First of all, the subtype of these ion channels could be identified as neuroendocrine subtype of L-type channels. Recent studies demonstrated that the neuroendocrine subtype of L-type channels is regulated by serine/ threonine kinases and protein tyrosine kinases. These phosphorylation-dependent regulatory mechanisms lead to Ca2+ fluxes into the cell which are independent of changes in the membrane potential and induced by a shift in the voltage-dependence ofthese ion channels. The regulation modality implied that L-type Ca2+ channels play an important role in signal transduction pathways which are important for a communication between retinal pigment epithelium and photoreceptors. L-type Ca2+ channels in the retinal pigment epithelium seem to be involved in the regulation of secretion of various factors, in growth factor-dependent intracellular signalling and in the regulation of the phagocytosis of photoreceptor outer membranes. Thus, voltage-dependent Ca2+ channels in the retinal pigment epithelium are of importance for the function of photoreceptors.

Animals↗