Transfection of myeloid leukaemia cell lines is distinctively regulated by fibronectin substratum

Gene transfer into haematopoietic cells is a challenging approach. The extracellular matrix component fibronectin has been known to modulate the cell cycle dynamics, viability and differentiation of leukaemia cells. Thus, our aim was to investigate the influence of fibronectin substratum on the liposomal transfection of myeloid leukaemia cell lines. Liposomal transfection was performed with K562 and HL-60 as representative lines of transfection-competent and -incompetent myeloid leukaemia cells, respectively. Flow cytometry analyses were performed to determine transfection efficiency monitored by green fluorescent protein (GFP) expression and to assess cell viability and cell cycle status. Quantitation of GFP gene expression and DNA uptake was assayed by real time PCR. The current data showed that the adhesion to fibronectin deteriorated the transfection of K562 cells. In contrary, it enhanced the delivery of plasmid DNA into HL-60 cells. Correspondingly, the adhesion to fibronectin influenced the transfection efficiency mainly by modulating the intracellular presence of plasmid DNA. The cell cycle and viability which is regulated by fibronectin had a minor impact on the success of gene delivery. This phenomenon may be considered as an important factor which may modulate the potential gene transfer approaches for myeloid leukaemia.     

Cardioprotective Effect of Selenium Against Cyclophosphamide-Induced Cardiotoxicity in Rats

The objective of this study is to evaluate the possible protective effects of selenium (Se) against cyclophosphamide (CP)-induced acute cardiotoxicity in rats. A total of 42 male Spraque-Dawley rats were divided into six groups (n = 7). Rats in the first group were served as control. Rats in the second group received CP (150 mg/kg) at the sixth day of experiment. Animals in the third and fourth groups were treated with only 0.5 and 1 mg/kg Se respectively for six consecutive days. Rats in the fifth and sixth groups received respective Se doses (0.5 or 1 mg/kg) for 6 days and then a single dose of CP administered on the sixth day. On day 7, the animals were sacrificed; blood samples were collected to measure malondialdehyde (MDA), glutathione (GSH), lactate dehydrogenase (LDH), creatine kinase-MB (CK-MB), and ischemia-modified albumin (IMA) levels. Heart tissues were processed routinely and tissue sections were stained with H + E for light microscopic examination. In the CP-treated rats MDA, LDH, CK-MB, and IMA serum levels increased, while GSH levels decreased. Microscopic evaluation showed that tissue damage was conspicuously lower in CP plus Se groups. Moreover, 1 mg/kg Se was more protective than 0.5 mg/kg Se as indicated by histopathological and biochemical values. In conclusion, Se is suggested to be a potential candidate to ameliorate CP-induced cardiotoxicity which may be related to its antioxidant activity.     

Relationships among Iron,Protein Oxidation and Lipid Peroxidation Levels in Rats with Alcohol-induced Acute Pancreatitis

It has been previously shown that alcohol induces the damage of pancreatic parenchyma tissue, but the mechanism of this damage is still poorly understood. Assuming that oxygen radical damage may be the involved, we measured markers of oxidative damage in pancreatic tissue, blood serum, plasma, and whole blood of rats with early-stage alcohol-induced acute pancreatitis. Thirty-eight male Wistar rats were divided into three groups: the control group (group 1), the acute pancreatitis group 1 day (group 2), and 3 days (group 3) after the injection of ethyl alcohol into the common biliary duct, respectively. The levels of Fe in tissue and serum, whole blood viscosity, plasma viscosity, fibrinogen and homocysteine (Hcy) levels, erythrocyte and plasma malondialdehyde (MDA), and tissue and plasma protein carbonyl levels were found to be significantly higher in groups 2 and 3 than in group 1. However, the levels of reduced glutathione (GSH) in tissue and erythrocytes were significantly lower in groups 2 and 3 than in group 1. These results suggest that elevated Fe levels in serum and pancreatic tissue in rats with early-stage alcohol-induced acute pancreatitis is associated with various hemorheological changes and with oxidative damage of the pancreas.     

Macrosomic newborns of diabetic mothers are associated with increased aortic intima-media thickness and lipid concentrations

BACKGROUND/AIMS: Exposure to diabetes in utero has been established as a significant risk factor for some of components of metabolic syndrome. A few studies have examined relationship between the metabolic syndrome and echocardiographic left ventricular (LV) mass. We aimed to investigate relationship between abdominal aortic intima-media thickness (aIMT), LV mass and lipid profile in macrosomic newborns. METHODS: Abdominal aIMT was measured in 30 macrosomic neonates of diabetic mothers (group A), 30 macrosomic neonates of healthy mothers (group B) and 30 healthy neonates (group C). Lipid profile and LV mass were determined. RESULT: Mean aIMT was significantly higher in groups A and B (0.489 +/- 0.015, 0.466 +/- 0.019 mm, respectively) than in controls (0.375 +/- 0.024 mm). Weight-adjusted aIMT in group A was significantly higher than in groups B and C. Macrosomia was associated with increased lipid concentrations. Both LV mass indexed for BSA (body surface area) and birth weight measurements were significantly increased in group A compared with control. CONCLUSIONS: Macrosomic neonates of diabetic mothers have significant aIMT and LV mass indexed for BSA and birth weight with lipid alterations. It might play a role in the pathogenesis of atherosclerosis in adult life.     

Protective Effect of Seleno-<Emphasis Type="SmallCaps">l</Emphasis>-Methionine on Cyclophosphamide-Induced Urinary Bladder Toxicity in Rats

Cyclophosphamide (CP) is a widely used antineoplastic drug, which could cause toxicity of the normal cells due to its toxic metabolites. Its urotoxicity may cause dose-limiting side effects like hemorrhagic cystitis. Overproduction of reactive oxygen species (ROS) during inflammation is one of the reasons of the urothelial injury. Selenoproteins play crucial roles in regulating ROS and redox status in nearly all tissues; therefore, in this study, the urotoxicity of CP and the possible protective effects of seleno-l-methionine (SLM) on urinary bladder of rats were investigated. Intraperitoneal (i.p.) administration of 50, 100, or 150 mg/kg CP induced cystitis, in a dose-dependent manner, as manifested by marked congestion, edema and extravasation in rat urinary bladder, a marked desquamative damage to the urothelium, severe inflammation in the lamina propria, focal erosions, and polymorphonuclear (PMN) leukocytes associated with occasional lymphocyte infiltration determined by macroscopic and histopathological examination. In rat urinary bladder tissue, a significant decrease in the endogenous antioxidant compound glutathione, and elevation of lipid peroxidation were also noted. Pretreatment with SLM (0.5 or 1 mg/kg) produced a significant decrease in the bladder edema and caused a marked decrease in vascular congestion and hemorrhage and a profound improvement in the histological structure. Moreover, SLM pretreatment decreased lipid peroxide significantly in urinary bladder tissue, and glutathione content was greatly restored. These results suggest that SLM offers protective effects against CP-induced urinary bladder toxicity and could be used as a protective agent against the drug toxicity.     

Dual actions of the antioxidant chlorophyllin,a glutathione transferase P1-1 inhibitor,in tumorigenesis and tumor progression

Glutathione (GSH) and enzymes related to this antioxidant molecule are often overexpressed in tumor cells and may contribute to drug resistance. Blockade of glutathione transferases (GSTs) has been proposed to potentiate the efficacy of chemotherapeutic drugs in cancer. The aim of this study was to evaluate the effect of chlorophyllin that has antioxidant properties, and also interferes with the activity of GST P1-1, on breast cancers in vitro and in vivo. The in vivo studies were conducted using an N-methyl- N-nitrosourea (MNU)-induced chemical carcinogenesis model in laboratory rats. DNA damage, GST activity, and GSH levels were determined in liver and tumor tissues. Treatment with chlorophyllin increased the GSH levels in the liver and significantly decreased DNA damage in the blood, liver, and tumor tissues. Even though tumorigenesis was delayed in rats receiving chlorophyllin before MNU injections, once the tumors emerged, the progression of tumor appeared to be faster than in the animals that received the carcinogen only. Out of nine breast cell lines, GST P1-1 expression was detected in MCF-12A, MDA-MB-231, and HCC38. Concomitant incubation with chlorophyllin and docetaxel did not significantly affect cell proliferation and viability. Chlorophyllin displayed genoprotective effects that initially delayed tumorigenesis. However, once the tumors were established, it may act as a promoter that facilitates tumor growth, potentially by a mechanism independent of cell proliferation and viability. Our results underline the pros and cons of antioxidant treatment in cancer, even if it has a capacity to inhibit GST P1-1.     

Risk factors for radiographic progression in psoriatic arthritis: subanalysis of the randomized controlled trial ADEPT

Introduction  

To identify independent predictors of radiographic progression in psoriatic arthritis (PsA) for patients treated with adalimumab or placebo in the Adalimumab Effectiveness in PsA Trial (ADEPT).     

Function of Ech Hydrogenase in Ferredoxin-Dependent,Membrane-Bound Electron Transport in Methanosarcina mazei

Reduced ferredoxin is an intermediate in the methylotrophic and aceticlastic pathway of methanogenesis and donates electrons to membrane-integral proteins, which transfer electrons to the heterodisulfide reductase. A ferredoxin interaction has been observed previously for the Ech hydrogenase. Here we present a detailed analysis of a Methanosarcina mazei Δech mutant which shows decreased ferredoxin-dependent membrane-bound electron transport activity, a lower growth rate, and faster substrate consumption. Evidence is presented that a second protein whose identity is unknown oxidizes reduced ferredoxin, indicating an involvement in methanogenesis from methylated C₁ compounds.The aceticlastic pathway of methanogenesis creates approximately 70% (10) of the biologically produced methane and is of great ecological importance, as methane is a potent greenhouse gas. Organisms using this pathway to convert acetate to methane belong exclusively to the genera Methanosarcina and Methanosaeta. The two carbon atoms of acetate have different fates in the pathway. The methyl moiety is converted to methane, whereas the carbonyl moiety is further oxidized to CO₂ and the electrons derived from this oxidation step are used to reduce ferredoxin (Fd) (6). During methanogenesis from methylated C₁ compounds (methanol and methylamines), one-quarter of the methyl groups are oxidized to obtain electrons for the reduction of heterodisulfide (27). A key enzyme in the oxidative part of methylotrophic methanogenesis is the formylmethanofuran dehydrogenase, which oxidizes the intermediate formylmethanofuran to CO₂ (7). The electrons are transferred to Fd. It has been suggested that reduced ferredoxin (Fd_red) donates electrons to the respiratory chain with the heterodisulfide (coenzyme M [CoM]-S-S-CoB) as the terminal electron acceptor and that the reaction is catalyzed by the Fd_red:CoM-S-S-CoB oxidoreductase system (7, 24). The direct membrane-bound electron acceptor for Fd_red is still a matter of debate; for the Ech hydrogenase, a reduced ferredoxin-accepting, H₂-evolving activity has been observed for Methanosarcina barkeri (20), which implies that the H₂:CoM-S-S-CoB oxidoreductase system is involved in electron transport (13). Direct electron flow from the Ech hydrogenase to the heterodisulfide reductase has not been shown to date (20, 21). In contrast to M. barkeri, Methanosarcina acetivorans lacks the Ech hydrogenase (11). It can nevertheless grow on acetate, which is why another complex present in this organism, the Rnf complex, is thought to be involved in the aceticlastic pathway of methanogenesis as an acceptor for Fd_red (8, 10, 17). The Methanosarcina mazei genome, however, contains genes coding for the Ech hydrogenase, but this species lacks the Rnf complex (5).To investigate whether the Ech hydrogenase is the only means by which M. mazei channels electrons from Fd_red into the respiratory chain, a mutant lacking the Ech hydrogenase (M. mazei Δech mutant) was constructed. Electron transport experiments using Fd_red as the electron donor and CoM-S-S-CoB as the electron acceptor were conducted with wild-type and mutant membranes to gain deeper insight into the actual membrane-bound protein complexes that accept electrons from Fd_red. Furthermore, an in-depth characterization of the growth and trimethylamine (TMA) consumption of the Δech mutant was performed, which provided insight into the in vivo role of Ech hydrogenase.     

Effect of drought stress implemented at pre- or post-anthesis stage on some physiological parameters as screening criteria in chickpea cultivars

Drought is one of the most important factors limiting chickpea production in arid and semi-arid regions. There is little information regarding genotypic variation for drought tolerance in chickpea cultivars. Screening for drought tolerance is very important. It is essential to identify the physiological mechanisms of drought tolerance to complete conventional breeding program. Glasshouse experiment was carried out to study the genotypic variation among 11 chickpea (Cicer arietinum L.) cultivars. Plants were grown either under optimum conditions or drought stress was implemented at pre-or post-anthesis stages. The drought susceptibility index (DSI) was used as the measure of drought tolerance. Relationships between DSI and excised-leaf water loss (RWL), relative water content (RWC), membrane permeability, ascorbic acid, proline, and chlorophyll contents, lipid peroxidation, and hydrogen peroxide concentrations were determined in order to find out whether these physiological parameters could be used as the genotypic selection criteria for drought tolerance. The results of this study indicated that there was a wide variation in tolerance to drought stress among the chickpea cultivars, which could be exploited in breeding new chickpea cultivars with high drought tolerance. The results also demonstrated that drought-tolerant cultivars had a higher RWC, ascorbic acid and proline concentrations, but lower RWL and membrane permeability in comparison to drought-sensitive cultivars. The significant and a well defined relationships between DSI and RWC, RWL, ascorbic acid, proline, and membrane permeability were found. It was concluded that these parameters could be instrumental in predicting the drought tolerance of chickpea cultivars. This text was submitted by the authors in English.