Magnetite Nanoparticles Inhibit Tumor Growth and Upregulate the Expression of P53/P16 in Ehrlich Solid Carcinoma Bearing Mice

Background

Magnetite nanoparticles (MNPs) have been widely used as contrast agents and have promising approaches in cancer treatment. In the present study we used Ehrlich solid carcinoma (ESC) bearing mice as a model to investigate MNPs antitumor activity, their effect on expression of p53 and p16 genes as an indicator for apoptotic induction in tumor tissues.

Method

MNPs coated with ascorbic acid (size: 25.0±5.0 nm) were synthesized by co-precipitation method and characterized. Ehrlich mice model were treated with MNPs using 60 mg/Kg day by day for 14 injections; intratumorally (IT) or intraperitoneally (IP). Tumor size, pathological changes and iron content in tumor and normal muscle tissues were assessed. We also assessed changes in expression levels of p53 and p16 genes in addition to p53 protein level by immunohistochemistry.

Results

Our results revealed that tumor growth was significantly reduced by IT and IP MNPs injection compared to untreated tumor. A significant increase in p53 and p16 mRNA expression was detected in Ehrlich solid tumors of IT and IP treated groups compared to untreated Ehrlich solid tumor. This increase was accompanied with increase in p53 protein expression. It is worth mentioning that no significant difference in expression of p53 and p16 could be detected between IT ESC and control group.

Conclusion

MNPs might be more effective in breast cancer treatment if injected intratumorally to be directed to the tumor tissues.     

Salarin C inhibits the maintenance of chronic myeloid leukemia progenitor cells

We previously showed that incubation of chronic myeloid leukemia (CML) cells in very low oxygen selects a cell subset where the oncogenetic BCR/Abl protein is suppressed and which is thereby refractory to tyrosine kinase inhibitors used for CML therapy. In this study, salarin C, an anticancer macrolide extracted from the Fascaplysinopsis sponge, was tested as for its activity on CML cells, especially after their incubation in atmosphere at 0.1% oxygen. Salarin C induced mitotic cycle arrest, apoptosis and DNA damage. Salarin C also concentration-dependently inhibited the maintenance of stem cell potential in cultures in low oxygen of either CML cell lines or primary cells. Surprisingly, the drug also concentration-dependently enforced the maintenance of BCR/Abl signaling in low oxygen, an effect which was paralleled by the rescue of sensitivity of stem cell potential to IM. These results suggest a potential use of salarin C for the suppression of CML cells refractory to tyrosine kinase inhibitors     

Prevalence of Diterpenes in Essential Oil of Euphorbia mauritanica L.: Detailed Chemical Profile,Antioxidant, Cytotoxic and Phytotoxic Activities

Plants belonging to Euphorbia L. genus are considered very interesting from a medicinal point of view due to their diverse metabolites and bioactivities. The essential oil (EO) of Euphorbia mauritanica L. is not studied up to date. Therefore, the present study aimed to explore the chemical profile of this EO and evaluate its antioxidant, cytotoxic, and allelopathic potentialities. The EO was extracted from the whole plant via hydrodistillation and then, analyzed by gas chromatography/mass spectrometry (GC/MS). The correlation of E. mauritanica with the other Euphorbia plants was established using chemometric analysis. The antioxidant activity was determined based on scavenging of the free radical, 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). The anti-proliferation of the EO on the Hep G2 and MCF-7 cells was evaluated. Finally the allelopathic activity of the EO was assessed against the two noxious weeds, Dactyloctenium aegyptium and Urospermum picroides. Forty-one compounds were identified using GC/MS analysis, with an abundance of terpenoids (91.54 %) that were categorized into mono- (30.75 %), sesqui- (15.23 %), and diterpenes (45.56 %). Interestingly, the results revealed the preponderance of diterpenoid constituents although they are rarely found in the EOs of the plant kingdom. The major compounds were (3E)-cembrene A (18.66 %), verticiol (17.05 %), limonene (7.91 %), eucalyptol (7.26 %), α-pinene (5.61 %), neo-cembrene A (3.52 %), kaur-16-ene (3.24 %), and cembrene (3.09 %). The EO showed moderate antioxidant activity where it attained IC₅₀ values of 83.34 and 64.21 μg mL⁻¹ for DPPH and ABTS compared to 23.01 and 19.23 μg mL⁻¹ for ascorbic acid as standard, respectively. The EO exhibited very weak cytotoxic effect on MCF-7 and Hep G2 cells. The EO showed significant allelopathic activities against the weeds D. aegyptium and U. picroides in a concentration-dependent manner. EO was found more effective against U. picroides than D. aegyptium with IC₅₀ values of 0.79, 0.45, and 0.67 mg mL⁻¹ and 1.17, 0.55, and 1.08 mg mL⁻¹ for germination, root, and shoot growth, respectively. Due to the high content of diterpenes in E. mauritanica, further study is recommended for more characterization of pure forms of the identified diterpenes as well as evaluating their bioactivity either solely or synergistically.     

The protective effect of melatonin in lungs of newborn rats exposed to maternal nicotine

We investigated possible healing effects of melatonin (MEL) on biochemical and histological changes in the lungs of rat offspring caused by exposure to nicotine (NT) in utero. Pregnant rats were divided randomly into five groups. The SP group was treated with physiological saline. The EA group was treated with ethyl alcohol. The MEL group was treated with MEL. The NT group was treated with NT. The NT + MEL group was treated with NT and MEL. At the end of the study, the biochemistry and histopathology of lung tissue of the offspring were examined. Reduced alveolar development and increased numbers of alveolar macrophages and mast cells were observed in the NT group compared to the SP, EA and MEL groups. We also found increased malondialdehyde (MDA) levels and decreased total glutathione (GSH) levels in the NT group. Application of MEL ameliorated the histological and biochemical damage caused by NT. The number of alveoli was greater in the NT + MEL group than in the NT group. Also, the increased numbers of alveolar macrophages and mast cells resulting from exposure to NT were decreased following MEL treatment. We found that MEL caused a significant decrease in the level of MDA. Maternal exposure to NT caused significant structural and biochemical changes in the lungs of the offspring and administration of MEL ameliorated the changes.     

New nitric oxide donating 1,2,4-triazole/oxime hybrids: Synthesis,investigation of anti-inflammatory,ulceroginic liability and antiproliferative activities

A series of novel nitric oxide (NO) donating triazole/oxime hybrids was prepared and evaluated for their anti-inflammatory activity. Most of the tested compounds showed significant anti-inflammatory activity using carrageenan-induced rat paw edema method compared to indomethacin. Calculation of the ulcer indices and histopathological investigation indicated that the prepared NO-donating oximes exhibited less ulcerogenicity compared to their intermediate ketones and indomethacin. The NO-donating oxime 6i revealed significant activity against renal cancer A498 cell lines with 50.52 cell growth inhibition.     

Drug-metabolizing enzymes and praziquantel bioavailability in mice harboring Schistosoma mansoni isolates of different drug susceptibilities

The level of drug-metabolizing enzymes (cytochrome P450 [CYP450] and cytochrome b5 [cyt b5]) and the bioavailability of praziquantel (PZQ) were investigated in batches of mice infected with Schistosoma mansoni displaying either a decreased susceptibility to PZQ ("EE2" and "BANL"-isolates), or a normal susceptibility to the drug ("CD" isolate). Each batch was divided into 2 groups. The first group was further subdivided into 5 subgroups. Subgroups 1 to 4 were treated 7 wk postinfection (PI) with oral PZQ at 25, 50, 100, and 200 mg/kg for 5 consecutive days, whereas the fifth subgroup was administered the vehicle only as control. Animals were perfused 9 wk PI, and worms were counted to estimate PZQ ED50. CYP450 and cyt b5 were examined in hepatic microsomes of infected untreated mice and of infected mice treated with 25 and 200 mg/ kg PZQ. The second group was given PZQ 7 wk PI and was further subdivided into 11 subgroups, killed at 2, 5, 15, 30, 60, 90, 120, 150, 180, 240, and 360 min postdosing to study pharmacokinetic parameters of PZQ. Mice harboring S. mansoni isolates having higher PZQ ED50 (170.3 mg/kg for EE2 and 249.9 mg/kg for BANL vs. 82.96 mg/kg for CD) had higher levels of CYP450 and cyt b5, a PZQ Cmax decreased by 19-30% and area under the serum concentration-time curve0-6 hr decreased by 57-74%. Data suggest that S. mansoni isolates that are less sensitive to PZQ induce a lower inhibition of hepatic drug-metabolizing enzymes, with a consequently higher metabolic transformation of PZQ.     

Radioprotective effect of Curcuma longa extract on γ-irradiation-induced oxidative stress in rats

This study was conducted to evaluate the modulatory effect of aqueous extract of Curcuma longa (L.) against γ-irradiation (GR), which induces biochemical disorders in male rats. The sublethal dose of GR was determined in primary hepatocytes. Also, the effect of C.?longa extract was examined for its activity against GR. In rats, C.?longa extract was administered daily (200?mg/kg body mass) for 21?days before, and 7?days after GR exposure (6.5 Gy). The lipid profile and antioxidant status, as well as levels of transaminases, interleukin-6 (IL-6), and tumour necrosis factor?α (TNFα) were assessed. The results showed that in hepatocytes, the aqueous extract exhibited radioprotective activity against exposure to GR. Exposure of untreated rats to GR resulted in transaminase disorders, lipid abnormalities, elevation of lipid peroxidation, trace element alterations, release of IL-6 and TNF, and decrease in glutathione and protein level of superoxide dismutase-1 (SOD-1) and peroxiredoxin-1 (PRDX-1). However, treatment of rats with this extract before and after GR exposure improved antioxidant status and minimized the radiation-induced increase in inflammatory cytokines. Changes occurred in the tissue levels of trace elements, and the protein levels of SOD-1 and PRDX-1 were also modulated by C.?longa extract. Overall, C.?longa exerted a beneficial radioprotective effect against radiation-induced oxidative stress in male rats by alleviating pathological disorders and modulating antioxidant enzymes.     

Synthesis of some pyrazolyl benzenesulfonamide derivatives as dual anti-inflammatory antimicrobial agents

Four series of pyrazolylbenzenesulfonamide derivatives were synthesized and evaluated for their anti-inflammatory activity using cotton pellet induced granuloma and carrageenan-induced rat paw edema bioassays. Moreover, COX-1 and COX-2 inhibitory activity, ulcerogenic effect and acute toxicity were also determined. Furthermore, the target compounds were screened for their in-vitro antimicrobial activity against Eischerichia coli, Staphylococcus aureus and Candida albicans. Compounds 4-(3-Phenyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9a and 4-(3-Tolyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9b were not only found to be the most active dual anti-inflammatory antimicrobial agents in the present study with good safety margin and minimal ulcerogenic effect but also exhibited good selective inhibitory activity towards COX-2. A docking pose for 9a and 9b separately in the active site of the human COX-2 enzyme was also obtained. Therefore, these compounds would represent a fruitful matrix for the development of dual anti-inflammatory antimicrobial candidates with remarkable COX-2 selectivity.     

The Association of the Arabidopsis Actin-Related Protein2/3 Complex with Cell Membranes Is Linked to Its Assembly Status But Not Its Activation

In growing plant cells, the combined activities of the cytoskeleton, endomembrane, and cell wall biosynthetic systems organize the cytoplasm and define the architecture and growth properties of the cell. These biosynthetic machineries efficiently synthesize, deliver, and recycle the raw materials that support cell expansion. The precise roles of the actin cytoskeleton in these processes are unclear. Certainly, bundles of actin filaments position organelles and are a substrate for long-distance intracellular transport, but the functional linkages between dynamic actin filament arrays and the cell growth machinery are poorly understood. The Arabidopsis (Arabidopsis thaliana) “distorted group” mutants have defined protein complexes that appear to generate and convert small GTPase signals into an Actin-Related Protein2/3 (ARP2/3)-dependent actin filament nucleation response. However, direct biochemical knowledge about Arabidopsis ARP2/3 and its cellular distribution is lacking. In this paper, we provide biochemical evidence for a plant ARP2/3. The plant complex utilizes a conserved assembly mechanism. ARPC4 is the most critical core subunit that controls the assembly and steady-state levels of the complex. ARP2/3 in other systems is believed to be mostly a soluble complex that is locally recruited and activated. Unexpectedly, we find that Arabidopsis ARP2/3 interacts strongly with cell membranes. Membrane binding is linked to complex assembly status and not to the extent to which it is activated. Mutant analyses implicate ARP2 as an important subunit for membrane association.In plant cells, the actin cytoskeleton forms an intricate network of polymers that organizes the cytoplasm and defines the long-distance intracellular trafficking patterns of the cell. The actin network is critical not only for tip-growing cells (for review, see Cole and Fowler, 2006; Lovy-Wheeler et al., 2007) but also during the coordinated cell expansion that occurs in cells that utilize a diffuse growth mechanism (for review, see Wasteneys and Galway, 2003; Smith and Oppenheimer, 2005). For example, the polarized diffuse growth of leaf trichomes is highly sensitized to actin cytoskeleton disruption (Mathur et al., 1999; Szymanski et al., 1999), and a recent analysis of Arabidopsis (Arabidopsis thaliana) ACTIN mutants revealed widespread cell swelling and isotropic expansion in numerous cell types in the root and shoot (Kandasamy et al., 2009). The actin network is dynamic. The array reorganizes during cell morphogenesis (Braun et al., 1999; Szymanski et al., 1999) and in response to endogenous (Lemichez et al., 2001) and external (Hardham et al., 2007) cues. A major research goal is to better understand not only how plant cells convert G-actin subunits to particular actin filament arrays but also how the actin network interacts with the cellular growth machinery during cell expansion.This is a difficult problem to solve, because in expanding vacuolated cells the actin array adopts numerous configurations and consists of dense meshworks of cortical actin filaments and bundles (Baluska et al., 2000), thick actin bundles that penetrate the central vacuole (Higaki et al., 2006), and meshworks of filaments and bundles that surround the nucleus and chloroplasts (Kandasamy and Meagher, 1999; Collings et al., 2000). The spatial relationships between these actin networks and localized cell expansion are not obvious. Certainly, the plasma membrane-cell wall interface is a critical location for the regulated delivery and fusion of vesicles containing cell wall polysaccharides. Frequent reports of localized domains of enriched cortical actin signal at regions of presumed localized cell expansion have led to the widely held view that the cortical actin array creates local tracks for vesicle-mediated secretion (for review, see Smith and Oppenheimer, 2005; Hussey et al., 2006). In one study, the dynamics of actin filaments were analyzed in living hypocotyl epidermal cells that utilize a diffuse growth mechanism (Staiger et al., 2009). In this case, individual actin filaments are very unstable and randomly oriented; therefore, the precise relationships between cortical F-actin, vesicle delivery, and cell shape change remain obscure. The best known function for the actin cytoskeleton is that of a track for myosin-dependent vesicle and organelle trafficking (Shimmen, 2007). The actin bundle network mediates the transport of cargo between endomembrane compartments (Geldner et al., 2001; Kim et al., 2005) and the long-distance actomyosin transport of a variety of organelles, including the Golgi (Nebenfuhr et al., 1999; Peremyslov et al., 2008; Prokhnevsky et al., 2008). Generation of distributed (Gutierrez et al., 2009; Timmers et al., 2009) and localized (Wightman and Turner, 2008) actin bundle networks appears to define early steps in the trafficking of Golgi-localized cellulose synthase complexes to the sites of primary and secondary wall synthesis, respectively.Plant cells employ diverse collections of G-actin-binding proteins, actin filament nucleators, and actin-bundling and cross-linking proteins to generate and remodel the F-actin network (for review, see Staiger and Blanchoin, 2006). One actin filament nucleator, termed the Actin-Related Protein2/3 (ARP2/3) complex, controls numerous aspects of plant morphogenesis and development. The vertebrate complex consists of the actin-related proteins ARP2 and ARP3 and five other unrelated proteins termed ARPC1 to ARPC5, in order of decreasing mass. ARP2/3 in isolation is inactive, but in the presence of proteins termed nucleation-promoting factors such as WAVE/SCAR (for WASP family Verprolin homologous/Suppressor of cAMP Repressor), ARP2/3 is converted into an efficient actin filament-nucleating machine (for review, see Higgs and Pollard, 2001; Welch and Mullins, 2002). In mammalian cells, ARP2/3 activities are linked to membrane dynamics. Keratocytes that crawl persistently on a solid substrate appear to use ARP2/3-generated dendritic actin filament networks at the leading edge to either drive or consolidate plasma membrane protrusion (Pollard and Borisy, 2003; Ji et al., 2008). In many vertebrate cell types, ARP2/3 has a strong punctate intracellular localization (Welch et al., 1997; Strasser et al., 2004), which could reflect hypothesized activities at the Golgi (Stamnes, 2002) or late endosomal (Fucini et al., 2002; Holtta-Vuori et al., 2005) compartment.Genetic studies in plants reveal nonessential but widespread functions for ARP2/3. In the moss Physcomitrella patens, the ARPC4 and ARPC1 subunit genes are critical during tip growth of protonemal filaments (Harries et al., 2005; Perroud and Quatrano, 2006). In Arabidopsis, loss of either ARP2/3 subunit gene or mutations in WAVE complex genes that positively regulate ARP2/3 cause complicated syndromes, including the loss of polarized diffuse growth throughout the shoot epidermis, defective cell-cell adhesion, and decreased hypocotyl elongation (for review, see Szymanski, 2005). Altered responses to exogenous Suc (Li et al., 2004; Zhang et al., 2008) and reduced root elongation (Dyachok et al., 2008) are also reported for wave and arp2/3 strains. In higher plants, the involvement of ARP2/3 in tip growth and root hair development is more subtle. In Lotus japonicus, mutation of NAP1 and PIR1, known positive regulators of ARP2/3 (Basu et al., 2004; Deeks et al., 2004; El-Assal et al., 2004a), causes incompletely penetrant root hair phenotypes, but in the presence of symbiotic bacteria, the mutants have defective infection threads and reduced root nodule formation. Arabidopsis arp2/3 mutants do not have obvious tip growth defects in pollen tubes or root hairs, but in the presence of GFP:TALIN (Mathur et al., 2003b) and in double mutant combinations with the actin-binding protein CAP1 (Deeks et al., 2007), the effects of ARP2/3 on root hair growth are unmasked.In Arabidopsis, the genetics of the positive regulation of ARP2/3 are well characterized and appear to occur solely through another heteromeric complex termed WAVE (Eden et al., 2002; for review, see Szymanski, 2005). The putative WAVE/SCAR complex contains five subunits, one of which is the ARP2/3 activator SCAR. Plant SCARs contain conserved N-terminal and C-terminal domains that mediate interactions with other WAVE complex proteins and ARP2/3 activation, respectively (Frank et al., 2004; Basu et al., 2005). In nonplant systems, the regulatory relationships between WAVE and ARP2/3 appear to vary between cell types and species (for review, see Bompard and Caron, 2004; Stradal and Scita, 2006). However, in Arabidopsis, double mutant analyses indicate that WAVE is the sole pathway for ARP2/3 activation and that all subunits positively regulate ARP2/3 (Deeks et al., 2004; Basu et al., 2005; Djakovic et al., 2006). SCAR quadruple mutants are indistinguishable from arp2/3 null plants (Zhang et al., 2008). In moss, BRICK1 and ARP2/3 mutants have similar phenotypes, suggesting conserved regulatory relationships between WAVE and ARP2/3 in the plant kingdom (Harries et al., 2005; Perroud and Quatrano, 2006, 2008).Despite extensive molecular genetic knowledge about the ARP2/3 pathway and the strong actin cytoskeleton and growth phenotypes of arp2/3 plants, there are few direct data on the existence of the plant complex and its cellular function. There are reports of ARP2/3 localization based on the behavior of individual subunits (Le et al., 2003). In some cases, the results are weakened by the unknown specificity of heterologous ARP2/3 antibodies (Van Gestel et al., 2003; Fiserova et al., 2006). A specific antibody was raised against Silvetia ARP2 (Hable and Kropf, 2005). In developing zygotes, rhizhoid emergence is an early and actin-dependent developmental event, and at this stage a broad subcortical cone of ARP2 signal extends from the nuclear envelope toward the rhizhoid apex (Hable and Kropf, 2005). Double labeling experiments detected considerable overlap between ARP2 and actin, but surprisingly, there was a broad cortical domain of putative organelle-associated distal ARP2 that did not overlap with actin. In tip-growing P. patens chloronema cells, ARPC4 also appears to be membrane associated and localizes to a broad subcortical apical zone (Perroud and Quatrano, 2006). For these localization and genetic studies that rely on individual ARP2/3 subunits, it is important to prove that a plant ARP2/3 complex exists to test for an association of the complex with endomembrane compartments.In this paper, we provide several lines of evidence for an evolutionarily conserved pathway for ARP2/3 complex assembly in plant cells. These studies are based in part on genetic and biochemical analyses of the putative ARP2/3 subunit gene ARPC4. We found that disruption of the ARPC4 gene caused catastrophic disassembly of the complex and an array of phenotypes that were indistinguishable from known arp2/3 mutants. Chromatography experiments clearly revealed that functional hemagglutinin (HA)-tagged ARPC4 and endogenous ARP3 subunits assemble fully into ARP2/3 complexes. Surprisingly, much of the cellular pool of the plant ARP2/3 complex is membrane associated. An analysis of an extensive collection of wave and arp2/3 mutants allowed us to conclude that the normal association with membranes depended on the presence of ARP2 and the assembly status of the complex but not on the existence of an active pool of ARP2/3 in the cell.     

The four hexamerin genes in the honey bee: structure,molecular evolution and function deduced from expression patterns in queens,workers and drones

Background  

Hexamerins are hemocyanin-derived proteins that have lost the ability to bind copper ions and transport oxygen; instead, they became storage proteins. The current study aimed to broaden our knowledge on the hexamerin genes found in the honey bee genome by exploring their structural characteristics, expression profiles, evolution, and functions in the life cycle of workers, drones and queens.