Inhibition of protein kinase CK2 with the clinical-grade small ATP-competitive compound CX-4945 or by RNA interference unveils its role in acute myeloid leukemia cell survival,p53-dependent apoptosis and daunorubicin-induced cytotoxicity

Background

The involvement of protein kinase CK2 in sustaining cancer cell survival could have implications also in the resistance to conventional and unconventional therapies. Moreover, CK2 role in blood tumors is rapidly emerging and this kinase has been recognized as a potential therapeutic target. Phase I clinical trials with the oral small ATP-competitive CK2 inhibitor CX-4945 are currently ongoing in solid tumors and multiple myeloma.

Methods

We have analyzed the expression of CK2 in acute myeloid leukemia and its function in cell growth and in the response to the chemotherapeutic agent daunorubicin We employed acute myeloid leukemia cell lines and primary blasts from patients grouped according to the European LeukemiaNet risk classification. Cell survival, apoptosis and sensitivity to daunorubicin were assessed by different means. p53-dependent CK2-inhibition-induced apoptosis was investigated in p53 wild-type and mutant cells.

Results

CK2α was found highly expressed in the majority of samples across the different acute myeloid leukemia prognostic subgroups as compared to normal CD34⁺ hematopoietic and bone marrow cells. Inhibition of CK2 with CX-4945, K27 or siRNAs caused a p53-dependent acute myeloid leukemia cell apoptosis. CK2 inhibition was associated with a synergistic increase of the cytotoxic effects of daunorubicin. Baseline and daunorubicin-induced STAT3 activation was hampered upon CK2 blockade.

Conclusions

These results suggest that CK2 is over expressed across the different acute myeloid leukemia subsets and acts as an important regulator of acute myeloid leukemia cell survival. CK2 negative regulation of the protein levels of tumor suppressor p53 and activation of the STAT3 anti-apoptotic pathway might antagonize apoptosis and could be involved in acute myeloid leukemia cell resistance to daunorubicin.

     

Fibrin as a scaffold for cardiac tissue engineering

Fibrin is a natural biopolymer with many interesting properties, such as biocompatibility, bioresorbability, ease of processing, ability to be tailored to modify the conditions of polymerization, and potential for incorporation of both cells and cell mediators. Moreover, the fibrin network has a nanometric fibrous structure, mimicking extracellular matrix, and it can also be used in autologous applications. Therefore, fibrin has found many applications in tissue engineering, combined with cells, growth factors, or drugs. Because a major limitation of cardiac cell therapy is low cell engraftment, the use of biodegradable scaffolds for specific homing and in situ cell retention is desirable. Thus, fibrin-based injectable cardiac tissue engineering may enhance cell therapy efficacy. Fibrin-based biomaterials can also be used for engineering heart valves or cardiac patches. The aim of this review is to show cardiac bioengineering uses of fibrin, both as a cell delivery vehicle and as an implantable biomaterial.     

Dinosaur Tracks as Paleogeographic Constraints: New Scenarios for the Cretaceous Geography of the Periadriatic Region

A really unexpected finding of sauropod and theropod footprints in southern Latium raises to four the number of the trampled levels recognized in central and southern Italy. After the recent findings in Latest Jurassic and Early, mid and Late Cretaceous carbonate platform deposits of the Periadriatic region, dinosaur footprints seem to provide very important paleogeographic constraints for reconstructing the geodynamic history of the Mediterranean area. The presence of a varied ichnoassociation makes acceptance of the current paleogeographic models concerning the relative and absolute position of the Laziale-Abruzzese-Campano and of Apulian-Dinaric domains during the Late Cretaceous more and more problematic. Dinosaur footprints, combined with other paleontological data, demonstrate that these areas were never completely pulled apart by deep seaways, while frequent or continuous links between them, and to southern and northern mainlands, probably persisted. These data also allowed us to improve our understanding of the timing of the Mesozoic plate motion in this segment of the Western Tethys.     

Enhanced Slow-Wave EEG Activity and Thermoregulatory Impairment following the Inhibition of the Lateral Hypothalamus in the Rat

Neurons within the lateral hypothalamus (LH) are thought to be able to evoke behavioural responses that are coordinated with an adequate level of autonomic activity. Recently, the acute pharmacological inhibition of LH has been shown to depress wakefulness and promote NREM sleep, while suppressing REM sleep. These effects have been suggested to be the consequence of the inhibition of specific neuronal populations within the LH, i.e. the orexin and the MCH neurons, respectively. However, the interpretation of these results is limited by the lack of quantitative analysis of the electroencephalographic (EEG) activity that is critical for the assessment of NREM sleep quality and the presence of aborted NREM-to-REM sleep transitions. Furthermore, the lack of evaluation of the autonomic and thermoregulatory effects of the treatment does not exclude the possibility that the wake-sleep changes are merely the consequence of the autonomic, in particular thermoregulatory, changes that may follow the inhibition of LH neurons. In the present study, the EEG and autonomic/thermoregulatory effects of a prolonged LH inhibition provoked by the repeated local delivery of the GABA_A agonist muscimol were studied in rats kept at thermoneutral (24°C) and at a low (10°C) ambient temperature (Ta), a condition which is known to depress sleep occurrence. Here we show that: 1) at both Tas, LH inhibition promoted a peculiar and sustained bout of NREM sleep characterized by an enhancement of slow-wave activity with no NREM-to-REM sleep transitions; 2) LH inhibition caused a marked transitory decrease in brain temperature at Ta 10°C, but not at Ta 24°C, suggesting that sleep changes induced by LH inhibition at thermoneutrality are not caused by a thermoregulatory impairment. These changes are far different from those observed after the short-term selective inhibition of either orexin or MCH neurons, suggesting that other LH neurons are involved in sleep-wake modulation.     

Carrier-mediated uptake and phloem systemy of a 350-Dalton chlorinated xenobiotic with an alpha-amino acid function

In a previous paper we have shown that epsilon-(phenoxyalkanecarboxylyl)-L-Lys conjugates are potent inhibitors of amino acid transport systems and that it is possible to modulate the uptake inhibition by hydrophobic or hydrophilic additions in the 4-position of the aromatic ring (J.F. Chollet, C. Delétage, M. Faucher, L. Miginiac, J.L. Bonnemain [1997] Biochem Biophys Acta 1336: 331-341). In this report we demonstrate that epsilon-(2,4-dichlorophenoxyacetyl)-L-Lys (2,4D-Lys), one of the largest molecules of the series and one of the most potent inhibitors, is a highly permeant conjugate. Uptake of 2,4D-Lys by broad bean (Vicia faba) leaf discs is mediated by an active carrier system (Km1 = 0.2 mM; Vmax1 = 2.4 nmol x cm(-2) x h(-1) at pH 5.0) complemented by an important diffusive component. Among the compounds tested (neutral, basic, and acidic amino acids, auxin, glutathione, and sugars), only the aromatic amino acids clearly compete with 2,4D-Lys. The conjugate accumulates in the vein network, is exported toward the growing organs, and exhibits a distribution pattern different from that of the herbicide moiety. However, over time 2,4D-Lys progressively splits into 2,4D and lysine. Analyses by high-performance liquid chromatography and liquid scintillation spectrometry of the phloem sap collected from the castor bean system, used as a systemy test, indicate decreasing capacities of 2,4D, 2,4D-Lys, and glyphosate, respectively, to move from the epidermis cell wall to the sieve element. Our results show that it is possible to design synthesis of large-size xenobiotics (approximately 350 D) with a lipophilic pole, exhibiting high mobility within the vascular system.     

Circulating miRNA‐375 as a potential novel biomarker for active Kaposi’s sarcoma in AIDS patients

The aim of this study was to identify circulating microRNAs (miRNAs) that could be used as biomarkers in patients at risk for or affected by AIDS‐Kaposi's sarcoma (KS). Screening of 377 miRNAs was performed using low‐density arrays in pooled plasma samples of 10 HIV/human herpesvirus 8 (HHV8)‐infected asymptomatic and 10 AIDS‐KS patients before and after successful combined antiretroviral therapy (cART). MiR‐375 was identified as a potential marker of active KS, being the most down‐regulated in AIDS‐KS patients after cART and the most up‐regulated in naïve AIDS‐KS patients compared to naïve asymptomatic subjects. Validation on individual plasma samples confirmed that miR‐375 levels were higher in AIDS‐KS compared to asymptomatic patients, decreased after cART‐induced remission in most AIDS‐KS patients and increased in patients with active KS. In asymptomatic patients miR‐375 was up‐regulated after cART in both screening and validation. Statistical analyses revealed an association between miR‐375 changes and CD4 cell counts, which could explain the discordant cases and the opposite trend between asymptomatic and AIDS‐KS patients. These data suggest that circulating miR‐375 might be a good indicator of active AIDS‐KS. Moreover, changes in miR‐375 levels may have a prognostic value in HIV/HHV8‐infected patients undergoing treatment. Further large‐scale validation is needed.     

Effects of Partial Replacement of Corn with Glycerin on Ruminal Fermentation in a Dual-Flow Continuous Culture System

The objective of this study was to evaluate the effects of partially replacing dry ground corn with glycerin on ruminal fermentation using a dual-flow continuous culture system. Six fermenters (1,223 ± 21 ml) were used in a replicated 3x3 Latin square arrangement with three periods of 10 d each, with 7 d for diet adaptation and 3 d for sample collections. All diets contained 75% concentrate and three dietary glycerin levels (0, 15, and 30% on dry matter basis), totaling six replicates per treatment. Fermenters were fed 72 g of dry matter/d equally divided in two meals/d, at 0800 and 2000 h. Solid and liquid dilution rates were adjusted daily to 5.5 and 11%/h, respectively. On d 8, 9, and 10, samples of 500 ml of solid and liquid digesta effluent were mixed, homogenized, and stored at -20°C. Subsamples of 10 ml were collected and preserved with 0.2 mL of a 50% H₂SO₄ solution for later determination of NH₃-N and volatile fatty acids. Microbial biomass was isolated from fermenters for chemical analysis at the end of each experimental period. Data were analyzed using the MIXED procedure in SAS with α = 0.05. Glycerin levels did not affect apparent digestibility of DM (P _Lin. = 0.13; P _Quad. = 0.40), OM (P _Lin. = 0.72; P _Quad. = 0.15), NDF (P _Lin. = 0.38; P _Quad. = 0.50) and ADF (P _Lin. = 0.91; P _Quad. = 0.18). Also, glycerin inclusion did not affect true digestibility of DM (P _Lin. = 0.35; P _Quad. = 0.48), and OM (P _Lin. = 0.08; P _Quad. = 0.19). Concentrations of propionate (P < 0.01) and total volatile fatty acids (P < 0.01) increased linearly and concentrations of acetate (P < 0.01), butyrate (P = 0.01), iso-valerate (P < 0.01), and total branched-chain volatile fatty acids, as well as the acetate: propionate ratio (P < 0.01) decreased with glycerin inclusion. Linear increases on NH₃-N concentration in digesta effluent (P < 0.01) and on NH₃-N flow (P < 0.01) were observed due to glycerin inclusion in the diets. Crude protein digestibility (P = 0.04) and microbial N flow (P = 0.04) were greater in the control treatment compared with the other treatments and responded quadratically with glycerin inclusion. Furthermore, the inclusion of glycerin linearly decreased (P = 0.02) non-ammonia N flow. Glycerin levels did not affect the flows of total N (P _Lin. = 0.79; P _Quad. = 0.35), and dietary N (P _Lin. = 0.99; P _Quad. = 0.07), as well as microbial efficiency (P _Lin. = 0.09; P _Quad. = 0.07). These results suggest that partially replacing dry ground corn with glycerin may change ruminal fermentation, by increasing total volatile fatty acids, and propionate concentration without affecting microbial efficiency, which may improve glucogenic potential of beef cattle diets.