Akt is upstream and MAPKs are downstream of NF-κB in paclitaxel-induced survival signaling events, which are down-regulated by curcumin contributing to their synergism

Paclitaxel is the most promising chemotherapeutic agent of plant origin despite its high cost and dose-limiting toxicity. Our earlier report has shown that cervical cancer cells can be sensitized by curcumin to paclitaxel-induced apoptosis through down-regulation of NF-κB and Akt. In the present study we have attempted to decipher the signaling pathways regulating the synergism of paclitaxel and curcumin. The study has clearly proved that Akt and NF-κB function successively in the sequence of paclitaxel induced signaling events where Akt is upstream of NF-κB. While inhibition of NF-κB led to complete inhibition of the synergism of paclitaxel and curcumin, inhibition of Akt brought about only partial reduction of the same, suggesting that, apart from Akt, there are other pathways induced by paclitaxel leading to NF-κB activation, which are also down-regulated by curcumin. Inactivation of NF-κB did not affect the activation of Akt and survivin, while that of Akt significantly inhibited NF-κB and completely inhibited up-regulation of survivin. Up-regulation of Cyclin-D1, Cox-2, XIAP and cIAP1 and phosphorylation of MAPKs, were completely inhibited on inactivation of NF-κB assigning a key regulatory role to NF-κB in the synergistic effect of paclitaxel and curcumin. While up-regulation of survivin by paclitaxel is regulated by Akt, independent of NF-κB, inactivation of neither Akt nor NF-κB produced any change in Bcl-2 level suggesting a distinct pathway for its action. As curcumin could effectively down-regulate all these survival signals induced by paclitaxel, we suggest it as a potent chemosensitizer to improve the therapeutic index of paclitaxel.     

Exploring the evolution of multicellularity in Saccharomyces cerevisiae under bacteria environment: An experimental phylogenetics approach

There have been over 25 independent unicellular to multicellular evolutionary transitions, which have been transformational in the complexity of life. All of these transitions likely occurred in communities numerically dominated by unicellular organisms, mostly bacteria. Hence, it is reasonable to expect that bacteria were involved in generating the ecological conditions that promoted the stability and proliferation of the first multicellular forms as protective units. In this study, we addressed this problem by analyzing the occurrence of multicellularity in an experimental phylogeny of yeasts (Sacharomyces cerevisiae) a model organism that is unicellular but can generate multicellular clusters under some conditions. We exposed a single ancestral population to periodic divergences, coevolving with a cocktail of environmental bacteria that were inoculated to the environment of the ancestor, and compared to a control (no bacteria). We quantified culturable microorganisms to the level of genera, finding up to 20 taxa (all bacteria) that competed with the yeasts during diversification. After 600 generations of coevolution, the yeasts produced two types of multicellular clusters: clonal and aggregative. Whereas clonal clusters were present in both treatments, aggregative clusters were only present under the bacteria treatment and showed significant phylogenetic signal. However, clonal clusters showed different properties if bacteria were present as follows: They were more abundant and significantly smaller than in the control. These results indicate that bacteria are important modulators of the occurrence of multicellularity, providing support to the idea that they generated the ecological conditions‐promoting multicellularity.     

Decreased repair of radiation-induced DNA double-strand breaks with cellular differentiation.

Although the majority of mammalian cells in situ are terminally differentiated, most DNA repair studies have used proliferating cells. In an attempt to understand better the relationship between differentiation and DNA repair, we have used the murine 3T3-T proadipocyte cell line. In this model system, proliferating (stem) cells undergo growth arrest (GD cells) and subsequently terminally differentiate into adipocytes when exposed to media containing platelet-depleted human plasma. Pulsed-field gel electrophoresis was used to evaluate the induction and repair of DNA double-strand breaks (DSBs) after ionizing radiation. The levels of radiation-induced DSBs in GD and terminally differentiated cells were similar, but in both cases greater than those found in stem cells at each radiation dose tested (0 to 40 Gy); these differences appear to be due to growth arrest in G1 phase. DNA DSBs were repaired with biphasic kinetics for each cell type. For terminally differentiated cells 25% of DNA DSBs remained unrejoined compared with < 10% for GD and stem cells after a repair time of 4 h. These data indicate that terminal differentiation of 3T3-T cells is associated with a reduction in the repair of ionizing radiation-induced DNA DSBs.     

A microassay for measurement of Fc function of human immunoglobulin preparations by using tetanus toxoid as antigen.

In order to minimize possible adverse reactions, the functional integrity of proteins in products derived from human plasma has to be unaffected by methods of preparation and storage conditions. Numerous biologically relevant functions of IgG, a major component of immunoglobulin for intravenous use preparations (IVIG), rely on the integrity of Fc fragments. Manufacturers are obliged to prove that Fc-mediated functions are maintained in IVIG preparations. The European Pharmacopoeia's monograph proposes a Rubella antigen-based test for Fc function of immunoglobulins. We present a modification of the proposed method achieved by using more convenient and readily available tetanus toxoid as an alternative antigen target and by adapting the procedure for the use on microtitre plates, thus greatly enhancing its feasibility and sample throughput. The test conditions were optimized so that batch-to-batch variability in tetanus antibody content did not influence the result. The precision of the test was within +/- 5%. By using this test, we compared Fc functionality of 9 commercial IVIG-7S preparations, which were prepared by using different virus inactivation/removal approaches. No significant differences between them have been found.     

Agricultural mosaics maintain significant flower and visiting insect biodiversity in a global hotspot

Agricultural expansion requires mitigation to conserve biodiversity and maintain functional interactions across the transformed mosaic. Conservation outside protected areas (PAs) means providing increased spatial opportunities for populations to survive over the long-term. Here we assess the capacity of agricultural mosaics for maintaining indigenous biodiversity against the benchmark levels in neighbouring PAs. We do this in three geographical areas in the major biodiversity hotspot, the Cape Floristic Region. We focus on two functionally related groups, indigenous flowers and their associated native insects, to assess the biodiversity value of certain components of agricultural mosaics. These components include agriculturally disturbed land, semi-transformed, disturbed land, and remnant patches of natural land, as well as adjacent PAs as reference sites. Overall, species richness across the agricultural mosaics outside PAs was similar to that within PAs for insects, although more variable for plants. Nevertheless, sites outside PAs still retained great diversity of flowering plants. Across the agricultural mosaic, sites shared 20–38 % of their insect species, and 12–28 % of plant species, indicating substantial species turnover. Each particular agricultural component retained species not sampled elsewhere in the landscape, indicating that habitats outside PAs can make a significant contribution to the compositional biodiversity of an area. Our findings are positive for biodiversity conservation across these agricultural mosaics, with these semi-natural components within the production landscape, in effect, extending the size of the PAs for many species. Conservation planning should recognize the value of such agricultural mosaics.     

Mechanistic evaluation of the signaling events regulating curcumin-mediated chemosensitization of breast cancer cells to 5-fluorouracil

5-Fluorouracil (5-FU) is the first rationally designed antimetabolite, which achieves its therapeutic efficacy through inhibition of the enzyme thymidylate synthase (TS), which is essential for the synthesis and repair of DNA. However, prolonged exposure to 5-FU induces TS overexpression, which leads to 5-FU resistance in cancer cells. Several studies have identified curcumin as a potent chemosensitizer against chemoresistance induced by various chemotherapeutic drugs. In this study, we report for the first time, with mechanism-based evidences, that curcumin can effectively chemosensitize breast cancer cells to 5-FU, thereby reducing the toxicity and drug resistance. We found that 10 μM 5-FU and 10 μM curcumin induces a synergistic cytotoxic effect in different breast cancer cells, independent of their receptor status, through the enhancement of apoptosis. Curcumin was found to sensitize the breast cancer cells to 5-FU through TS-dependent downregulation of nuclear factor-κB (NF-κB), and this observation was confirmed by silencing TS and inactivating NF-κB, both of which reduced the chemosensitizing efficacy of curcumin. Silencing of TS suppressed 5-FU-induced NF-κB activation, whereas inactivation of NF-κB did not affect 5-FU-induced TS upregulation, confirming that TS is upstream of NF-κB and regulates the activation of NF-κB in 5-FU-induced signaling pathway. Although Akt/PI3kinase and mitogen-activated protein kinase pathways are activated by 5-FU and downregulated by curcumin, they do not have any role in regulating the synergism. As curcumin is a pharmacologically safe and cost-effective compound, its use in combination with 5-FU may improve the therapeutic index of 5-FU, if corroborated by in vivo studies and clinical trials.     

Amylocyclicin,a Novel Circular Bacteriocin Produced by Bacillus amyloliquefaciens FZB42

Bacillus amyloliquefaciens FZB42 is a Gram-positive plant growth-promoting bacterium with an impressive capacity to synthesize nonribosomal secondary metabolites with antimicrobial activity. Here we report on a novel circular bacteriocin which is ribosomally synthesized by FZB42. The compound displayed high antibacterial activity against closely related Gram-positive bacteria. Transposon mutagenesis and subsequent site-specific mutagenesis combined with matrix-assisted laser desorption ionization–time of flight mass spectroscopy revealed that a cluster of six genes covering 4,490 bp was responsible for the production, modification, and export of and immunity to an antibacterial compound, here designated amylocyclicin, with a molecular mass of 6,381 Da. Peptide sequencing of the fragments obtained after tryptic digestion of the purified peptide revealed posttranslational cleavage of an N-terminal extension and head-to-tail circularization of the novel bacteriocin. Homology to other putative circular bacteriocins in related bacteria let us assume that this type of peptide is widespread among the Bacillus/Paenibacillus taxon.     

Development and growth of immature hips

The authors investigated the development of the bony acetabulum in type IIa+ immature hips using ultrasound follow-up evaluation of the alpha and beta angle. The study comprised 900 hips in one-month old infants. In the initial ultrasound examination the alpha angle measured less than 60 degrees and the beta angle more than 55 degrees. In the second ultrasound examination at the age of two months a discrepancy between the bony and cartilaginous acetabular component was found in 15% of the hips, which means that the type IIa+ hips were transformed into the type IIa-. The infants with type IIa- hips had more than two risk factors for developmental dysplasia of the hip. The statistical relative risk was 17. These hips were treated with the Pavlik harness during two months on average. After treatment all hips had normal values the alpha and beta angles (more than 60 degrees and less than 55 degrees respectively). This study showed that each newborn and infant with a sonographically confirmed hip development disorder has to be included in the follow-up evaluation of acetabular development, which makes it possible to choose the most adequate therapeutic and prophylactic measures.     

Youth and AIDS--a study of attitudes,knowledge, behavior and risks in the post-war Croatia

According to the latest reports, the Eastern Europe currently exhibits the greatest relative increase in the number of newly registered HIV infections in the world. At the same time, Central Europe remains relatively spared from the epidemic, with reported rates significantly lower than those in both Eastern and Western Europe. Croatia geographically affiliates to Central Europe, but it has two specific potential risk factors in comparison to neighboring countries: recent War events and a summer season when immigration of large number of tourists from Central and Eastern Europe is expected. Therefore, it is critical to examine AIDS attitudes in young people, increase their knowledge, monitor their behavior and warn on risks in order to prevent larger spread of epidemics from Eastern Europe to Croatia. In this study, we report on a large related survey and education program among 17-year-old high school pupils that was conducted in years immediately following the War (1996-1999).     

Primary Ewing's sarcoma of the kidney: a case report

Primary adult Ewing's sarcoma is a rare entity. They most commonly occur in children and young adults. 6% of them are localized extraosseously. We present a case of a 51 year old patient with primary renal Ewing's sarcoma and multiple metastases in liver and iliac bone. Patients with metastatic disease are usually treated with aggressive chemotherapy and have a poor outcome. Our patient underwent complete surgical excision of tumour, and was treated with aggressive chemotherapy, respectively. Two and half years after presentation he is well, without any symptoms.