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Air- and water-stable rhenium carbonyl complexes are investigated as CO-carriers for studying the light-dependent release of small amounts of carbon monoxide under physiological conditions. The reported very low quantum yields are in a suitable range for studying potential applications as photochemical CO-releasing molecules (Photo-CORMs) without rapidly reaching cytotoxic levels of carbon monoxide. Preliminary results on bio-compatibility and selective uptake of the compounds into human colon carcinoma cells (Caco-2 cell line) are described.

The role of the mitogen-activated protein (MAP) kinase phosphatases (MKPs) in light-damaged cells is unclear. Therefore we investigated the involvement of MKP-1 in the regulation of apoptosis and cell survival mediated by MAP kinase pathways in light-damaged human retinal pigment epithelial cells (ARPE-19). Light dose-dependent changes in the expression of MKP-1 and in the phosphorylation status of the MAP kinases, c-Jun-N-terminal kinase (JNK) and p38 were demonstrated. Low light doses up to 2 J cm(-2) led to an upregulation of MKP-1 which resulted in the prevention of cell death by inactivating JNK kinase. However, higher light doses (> or =3 J cm(-2)) significantly reduced MKP-1 protein expression and subsequently led to an increased JNK kinase activity followed by a significant increase in cell death. JNK kinase inactivation by the JNK inhibitor SP600125 significantly reduced light-induced cell death, suggesting that the cytoprotective properties of MKP-1 are mediated mainly by the JNK MAP kinase pathway. Physiological concentrations of ascorbic acid or taurine were seen to prevent apoptosis and cell death in light-damaged ARPE-19 cells by reducing oxidative stress within cells, thus maintaining MKP-1 at high levels, leading to an inactivation of the JNK kinase pathway which resulted in an increased cell viability.

Context: With regard to two-dimensional (2D) cell cultures in flat dishes, most of the three-dimensional (3D) cell cultures have advantages and gain importance in life sciences. But, their higher complexity requires adaptation of existing methods or different measurement systems in order to achieve accurate results. Bio-Impedance spectroscopy (Bio-IS) is a label- free and non-destructive method that is considered for analysis of 3D cell models. Purpose: We design a new Bio-IS measurement system for a 3D liver cell model to assess cell seeding and drug effects. Experiment Methods: We measured reflection coefficient S11 with differently (disc- and needle-like) shaped electrodes, which are in defined contact with the 3D liver cell model, metabolic activity (MTT test), and cytotoxicity (LDH assay). Results: We demonstrate that reflection coefficient S11 may be utilized to assess the 3D liver cell system in the frequency range of β-dispersion at 73 MHz only by means of the needle-like electrodes. This enables a reliable label-free and non-destructive determination of cell seeding and drug effects in the 3D liver cell culture model. The physical results have been verified by destructive biochemical methods (LDH and MTT assay). Conclusion: Our Bio-IS system for 3D liver cell models using needle-like electrodes enables label-ree and nondestructive determination of cell seeding and drug effects.

permeability of the small molecules Lucifer Yellow (ca. 520 Da) and FITC-dextran (3– 5k Da) but not of large molecules (40 kDa FITC-dextran). The tight junction protein ZO-1 and its tyrosine phosphorylation were upregulated after 24 h treatment with APAP. Conclusions Tight junction proteins play an important role in maintaining the intestinal barrier function. We assume that APAP affects the paracellular transport pathway through disassembly of the tight junction. We suppose that APAP leads to remodelling of the tight junction protein ZO-1 through changes in the level of protein expression and in tyrosine phosphorylation. APAP may reduce the bioavailability of co-administered substances through remodelling the tight junction by regulating ZO-1 and its tyrosine phosphorylation. As a result, the paracellular transport activity for small molecules is decreased as is net intestinal absorption.

Proteasome inhibitors such as MG-132 are considered as potential therapeutical tools in different clinical settings. The dual specificity MAP-kinase phosphatase MKP-1 plays a role in balancing signals mediating cell death or survival. Here the effect of cell hydration on MG-132-induced MKP-1 expression was investigated in H4IIE rat hepatoma cells.Hyperosmolarity (405mosmol/l) increased MKP-1 expression by MG-132, which was accompanied by an induction of c-Fos, c-Jun, cJun Ser73 phosphorylation, and AP-1 DNA binding. MKP-1 induction by MG-132 plus hyperosmolarity was sensitive to inhibition of p38(MAPK) and c-Jun-N-terminal kinases (JNKs) but not extracellular signal-regulated kinases Erk-1/Erk-2, and was accompanied by a decline of MAP-kinase activities. Although hyperosmolarity increased overall protein ubiquitination in presence of MG-132, ubiquitination of MKP-1 was found under normo-, but not hyperosmotic conditions. Hyperosmolarity also enabled MG-132 to induce poly-ADP-ribose polymerase (PARP) cleavage which was sensitive to inhibition of p38(MAPK) and JNKs but not Erk-1/Erk-2. PARP cleavage and caspase-3 activation in H4IIE cells treated with hyperosmolarity plus MG-132 was further increased by vanadate, consistent with a contribution of MKP-1 to counterbalance proapoptotic MAP-kinase signals.The findings suggest that among other factors cell hydration critically determines the cellular response to proteasome inhibitors.

A contribution of intracellular dehydration to insulin resistance has been established in human subjects and in different experimental systems. Here the effect of hyperosmolarity (405 mosmol/l) on insulin-induced mitogen-activated protein (MAP) kinase phosphatase (MKP)-1 expression was studied in H4IIE rat hepatoma cells. Insulin induces robust MKP-1 expression which correlates with a vanadate-sensitive decay of extracellular-signal-regulated kinase (Erk-1/Erk-2) activity. Hyperosmolarity delays MKP-1 accumulation by insulin and this corresponds to impaired MKP-1 synthesis, whereas MKP-1 degradation remains unaffected by hyperosmolarity. Rapamycin, which inhibits signalling downstream from the mammalian target of rapamycin (mTOR) and a peptide inhibiting protein kinase C (PKC) zeta/lambda abolish insulin-induced MKP-1 protein but not mRNA expression, suggesting the involvement of the p70 ribosomal S6 protein kinase (p70S6-kinase) and/or the eukaryotic initiation factor 4E-binding proteins (4E-BPs) as well as atypical PKCs in MKP-1 translation. Hyperosmolarity induces sustained suppression of p70S6-kinase and 4E-BP1 hyperphosphorylation by insulin, whereas insulin-induced tyrosine phosphorylation of the insulin receptor (IR) beta subunit and the IR substrates IRS1 and IRS2, recruitment of the phosphoinositide 3-kinase (PI 3-kinase) regulatory subunit p85 to the receptor substrates as well as PI 3-kinase activation, and Ser-473 phosphorylation of protein kinase B and Thr-410/403 phosphorylation of PKC zeta/lambda are largely unaffected under hyperosmotic conditions. The hyperosmotic impairment of both, MKP-1 expression and p70S6-kinase hyperphosphorylation by insulin is insensitive to K(2)CrO(4), calyculin A and vanadate, and inhibition of the Erk-1/Erk-2 and p38 pathways. The suppression of MKP-1 may further contribute to insulin resistance under dehydrating conditions by allowing unbalanced MAP kinase activation.

Little is known about the regulation of signal transducer and activator of transcription (STAT) stability. Here the osmolarity-dependence of STAT3 stability, ubiquitination, Tyr(705) phosphorylation, STAT3 transactivation and gamma-fibrinogen (gamma-FBG) expression was studied in hepatoma cells. Hyper-osmolarity accelerated STAT3 degradation which was prevented by proteasome inhibitors. Hypo-osmolarity stabilized STAT3, most likely due to a decrease in STAT3 ubiquitination. Accordingly, STAT3 Tyr(705) phosphorylation, alpha(2)-macroglobulin promoter activity and gamma-FBG expression were osmosensitive. Modulation of STAT3 stability may contribute to a hydration dependence of acute phase protein expression.