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Intracranial hypertension (ICP) and visual impairment intracranial pressure (VIIP) are some of the consequences of long-term space missions. Here we examined the behavior of oligodendrocyte progenitors (OLPs) after space flight using time-lapse microscopy. We show that most OLPs divided more than ground control (GC) counterparts did. Nonetheless, a subpopulation of OLPs flown to space presented a significant increase in autophagic cell death. Examination of the proteomic profile of the secretome of space flown OLPs (SPC-OLPs) revealed that the stress protein heat shock protein-90 beta “HSP-90β” was the 5th most enriched (6.8 times) and the secreted protein acidic and rich in cysteine “SPARC” was the 7th most enriched (5.2 times), with respect to ground control cells. SPARC induces endoplasmic reticulum stress, which leads to autophagy. Given the roles and importance of these two proteins in mammalian cells’ metabolism, their upregulation may hold the key to modulating cell proliferation and autophagy, in order to mitigate ICP and VIIP during and after space missions.

Oxygen consumption is particularly elevated in cardiac cells as they are equipped with a large number of mitochondria and high levels of respiratory chain components. Consequently, production of reactive oxygen species (ROS) is tightly controlled as an imbalance in redox reactions can lead to irreversible cellular damage. siRNA-mediated down-regulation of protein kinase CK2 has been implicated in the accumulation of ROS in cells. The present study was undertaken in order to investigate the role of CK2 in redox homeostasis in cardiomyoblasts. We found that inhibition or silencing of CK2 causes elevated levels of ROS, notably superoxide radical, and this is accompanied by suppression of NF-κB transcriptional activity and mitochondrial dysfunction. We show that CK2 regulates the expression of manganese superoxide dismutase, the enzyme catalyzing the dismutation of superoxide, in cancer cells but not in cardiomyoblasts. Furthermore, we report evidence that impaired expression of CK2 results in destabilization of the Bcl-2 mammalian homolog Bcl-xL, which is known to stabilize the mitochondrial membrane potential, through a mechanism involving disruption of the chaperone function of heat shock protein 90. Analysis of differential mRNA expression related to oxidative stress revealed that CK2 silencing caused a statistically significant deregulation of four genes associated with the oxidative damage, i.e., Fmo2, Ptgs1, Dhcr24, and Ptgs2. Overall, the results reported here are consistent with the notion that CK2 plays a role in conferring protection against oxidative stress by positively regulating pro-survival signaling molecules and the protein folding machinery in cardiomyoblasts.

Background Vitamin D3 and its analogues have recently been shown to enhance the anti-tumour effects of 5- Fluorouracil (5-FU) both in vitro and in xenograft mouse model of colon cancer. This study measured the potential mechanism(s) by which vitamin D3 could synergise the tumouricidal activities of 5-FU in azoxymethane (AOM) rat model of colon cancer. Methods Seventy-five male Wistar rats were divided equally into 5 groups: Control, AOM, AOM-treated by 5-FU (5-FU), AOM-treated by vitamin D3 (VitD3), and AOM-treated by 5-FU + vitamin D3 (5-FU/D). The study duration was 15 weeks. AOM was injected subcutaneously for 2 weeks (15 mg/kg/week). 5-FU was injected intraperitoneally in the 9th and 10th weeks post AOM (8 total injections were given: 12 mg/kg/day for 4 successive days, then 6 mg/kg every other day for another 4 doses) and oral vitamin D3 (500 IU/rat/day; 3 days/week) was given from week 7 post AOM till the last week of the study. The colons were collected following euthanasia for gross and histopathological examination. The expression of β-catenin, transforming growth factor-β1 (TGF-β1), TGF-β type 2 receptor (TGF-βR2), smad4, inducible nitric oxide synthase (iNOS), and heat shock protein-90 (HSP-90) proteins was measured by immunohistochemistry. In colonic tissue homogenates, quantitative RT-PCR was used to measure the mRNA expression of Wnt, β-catenin, Dickkopf-1 (DKK-1) and cyclooxygenase-2 (COX-2) genes, while ELISA was used to measure the concentrations of TGF-β1, HSP-90 and COX-2 proteins. Results Monotherapy with 5-FU or vitamin D3 significantly decreased the number of grown tumours induced by AOM (P

Die Behandlung von metastasierten gastroenteropankreatischen neuroendokrinen Tumoren (GEP-NETs) ist weiterhin eine große klinische Herausforderung. Zusätzliche medikamentöse Interventionsmöglicheiten werden dringend benötigt. Die vermehrte Aktivierung des intrazellulären Phosphatidylinsositol-3-kinase (PI3K)-AKT Signalweg führt zu einem vermehrten Tumorwachstum und verhindert Apoptose bei verchiedenen Tumorarten. Weiterhin spielt der Signalweg eine wichtige Rolle in der Resistenzentwicklung gegenüber Zytostatika sowie bei der Angiogenese. Das Hitzeschockprotein 90 (HSP90) ist ein wichtiger Regulator von AKT. Außerdem konnte bei vielen Tumoren eine Überexpression von HSP90 gezeigt werden. Auch hat die vermehrte Expression von HSP90 in Tumoren eine wichtige prognostische Bedeutung. Mittels Echtzeit-Proliferations-Assays untersuchten wir die Wirkung von Triciribine, einem AKT-Inhibitor, und IPI504, ein Inhibitor von HSP90, auf drei verschiedene gastroenteropankreatische neuroendokrine Tumorzelllinien. Wir analysierten die Induktion von Apoptose und die Effekte der Substanzen auf verschiedene krebsassoziierte Genprodukte. Weiterhin untersuchten wir die Rolle des Enzymes PTEN als ein möglicher Prediktor für die Wirksamkeit einer AKT-Inhibition. Mögliche additive antineoplastische Effekte durch Kombination mit zytostatischen oder anderen enzymspezifischen Inhibitoren wurden untersucht. Auch die in vivo Effekte wurden mittels einer Hühnermembranuntersuchung analysiert. Dabei zeigte sowohl die Inhibition von AKT, als auch die Hemmung von HSP90 einen antiproliferativen Effekt bei gastroenteropankreatischen neuroendokrinen Tumoren. Die Kombination mit klassischen Zytostatika, also auch die Kombination mit anderen signalwegspezifischen Substanzen zeigten synergistische antiproliferative Effekte. In vivo Versuche bestätigten die antiproliferative Wirkung der beiden Substanzen. Die Expression von PTEN konnte als Prädiktor bezüglich der Sensitivität von GEP-NET-Zellen gegenüber einer AKT-Inhibition nachgewiesen werden. Zusammenfassend konnten wir zeigen, dass sowohl die AKT-Inhibition, als auch die HSP90-Inhibition in der Lage ist, das Wachstum von gastroenteropankreatischen neuroendokrinen Tumorzellen zu hemmen., Treatment of gastroenteropancreatic neuroendocrine tumors (GEP-NET) is still unsatisfactory and innovative thera¬peutic approaches are urgently needed. Up- regulation of phosphatidylinositol-3-kinase (PI3K)-AKT signaling facilitates tumor cell growth and inhibits cell demise. The AKT-pathway also plays an important role in cytostatic therapy resistance and response to hypoxia and angiogenesis. The Heat shock protein 90 (Hsp90) is an important regulator of AKT-activity. Furthermore, it is overexpressed in a wide range of tumors and an emerging target for the treatment of cancer. We therefore examined the potency of triciribine, an AKT inhibitor, and IPI504, a HSP90 inhibitor, in three distinct neuroendocrine gastrointestinal tumor cell lines, using real- time cell proliferation assays. Also, we investigated the induction of apoptosis and effects on a broad range of cancer-associated gene products. Furthermore, we characterized the role of PTEN as a possible predictor of sensitivity to triciribine in GEP-NETs. We looked for additive anti-neoplastic effects of the drugs when combined with conventional cytostatic drugs or other targeted drugs, affecting different molecules of the PI3K-AKT-pathway. We also assessed the potency of AKT- and HSP90 inhibition to slow tumor growth in vivo, by using the chick chorioallantoic membrane assay. Both, AKT- and HSP90 inhibition significantly suppressed growth in different neuroendocrine cell lines. Combinations with classic cytostatic drugs as well as drugs targeting other molecules of the PI3K-AKT-pathway led to synergistic anti-proliferative effects. Additional in vivo-evaluations confirmed the anti-neoplastic potency of triciribine and IPI504. Expression of PTEN was shown to predict sensitivity of cells towards AKT inhibition. Thus, our data show that inhibition of the AKT-pathway as well as inhibition of HSP90 potently reduces the growth of GEP- NET cells alone or in combination therapies.

Heat-shock protein 90 (Hsp90) coordinates the trafficking and regulation of diverse signalling proteins, but its precise role in regulating specific cellular targets is not known1,2. Here we show that Hsp90 associates with endothelial nitric oxide synthase (eNOS) and is rapidly recruited to the eNOS complex by agonists that stimulate production of nitric oxide, namely vascular endothelial growth factor, histamine and fluid shear stress. Moreover, the binding of Hsp90 to eNOS enhances the activation of eNOS. Inhibition of signalling through Hsp90 attenuates both agonist-stimulated production of nitric oxide and endothelium-dependent relaxation of isolated blood vessels. Our results indicate that Hsp90 facilitates signalling mediated by growth-factor, G-protein and mechanotransduction pathways that lead to the activation of eNOS. These observations indicate that in addition to its role as a molecular chaperone involved in protein folding and maturation, Hsp90 may also be recruited to cellular targets depending on the activation state of the cell.

We have recently demonstrated that excessive aldosterone (Aldo) secretion in primary aldosteronism (PA) is associated with red blood cells (RBC) senescence. These alterations were prevented/inhibited by cortisol (Cort) or canrenone (Can) raising the hypothesis that Aldo effects in RBC may be mediated by mineralocorticoid receptor (MR), though to date MR has never been demonstrated in human RBC. The aim of this multicenter comparative study was to investigate whether Aldo effects were mediated by MR in these a-nucleated cells. We included 12 healthy controls (HC) and 22 patients with PA. MR presence and activation were evaluated in RBC cytosol by glycerol gradient sedimentation, Western blotting, immuno-precipitation and radioimmunoassay. We demonstrated that RBC contained cytosolic MR, aggregated with HSP90 and other proteins to form multiprotein complex. Aldo induced MR to release from the complex and to form MR dimers which were quickly proteolyzed. Cort induced MR release but not dimers formation while Can was not able to induce MR release. In addition, RBC cytosol from PA patients contained significantly higher amounts of both MR fragments (p