Anti-Plasmodial Assessment of Four Different Iranian Propolis Extracts.

BACKGROUND: Eradication of malaria will depend on discovery of new intervention tools such as anti-malarial drugs. Due to the increasing interest in the application of propolis against significant clinical pathogenic agents, the aim of the present investigation was to evaluate the anti-plasmodial effect of Iranian propolis extracts against chloroquine (CQ)-sensitive Plasmodium falciparum 3D7 and Plasmodium berghei (ANKA strain). METHODS: Crude samples of honeybee (Apis mellifera) propolis were collected from four provinces in northern (Kalaleh, Golestan), northeastern (Chenaran, Razavi Khorasan), central (Taleghan, Alborz) and western (Morad Beyg, Hamedan) areas of Iran with different types of flora. The dried propolis samples were extracted with three different solvents, including ethanol 70% (EtOH), ethyl acetate (EA) and dichloromethane (DCM). RESULTS: All extracts were shown to have in vitro anti-plasmodial activity with IC50 ranging from 16.263 to 80.012 µg/mL using parasite lactate dehydrogenase (pLDH) assay. The DCM extract of Morad Beyg propolis indicated the highest anti-plasmodial activity (IC50: 16.263 ± 2.910 µg/mL; P = 0.027, Kruskal-Wallis H-test). The samples were also evaluated in mice for their in vivo anti-plasmodial effect. The curative effect against established infection (Rane test) showed that both extracts at all doses (50, 100, and 200 mg/kgBW) produced anti-plasmodial activity against the parasite. Furthermore, using gas chromatography-mass spectrometry (GC-MS), the quantity of flavonoids in DCM and EtOH 70% extracts were found to be 7.42% and 3.10%, respectively. CONCLUSION: The potent anti-plasmodial activity of both EtOH 70% and DCM extracts of the propolis of Morad Beyg, Hamedan suggests further analyses of individual components to assess its utilization as anti-malarial drugs.

Frequency of PTEN alterations, TMPRSS2-ERG fusion and their association in prostate cancer.

BACKGROUND: Phosphatase and tensin homolog (PTEN) gene aberration and trans membrane protease, serine 2 (TMPRSS2)-v-ets avian erythroblastosis virus E26 oncogene homolog (ERG) fusion are the most prevalent genomic events in prostate cancer. In this study we aimed to evaluate the frequency of PTEN alteration and TMPRSS2-ERG fusion and possible link between these two biomarkers in Iranian men. METHODS: We assessed 42 fresh frozen tissue samples of prostate cancer (PCA) obtained by radical prostatectomy, interrogating the TMPRSS2-ERG fusion gene along with PTEN gene status using Real Time PCR and FISH methods. RESULTS: Using Real Time PCR we identified the TMPRSS2-ERG fusion in 64% (27/42) of tumor samples, which was confirmed by FISH technique, giving 21 positive samples with deletion, suggesting the presence of TMPRSS2-ERG fusion gene. By contrast, PTEN deletion was detected in 52% (11/21) of PCA samples, which all showed low expression in Real Time. Concomitance of PTEN deletion or low expression and TMPRSS2-ERG fusion was present in PCA samples (P=0.005). All of the PTEN deletion samples showed TMPRSS2-ERG fusion, (11/11, 100%) while not all of the TMPRSS2-ERG fusion positive samples showed PTEN deletion. None of 29 cases of BPH and 8 cases of normal zone of tumor tissue showed TMPRSS2-ERG fusion. CONCLUSIONS: These results indicate that PTEN loss occurs in cooperation with TMPRSS2-ERG fusion in PCA. While the majority of PCA samples harbor TMPRSS2-ERG fusion as well as PTEN gene deletion, normal tissues do not show these molecular aberrations.

Stability and biological activity evaluations of PEGylated human basic fibroblast growth factor.

BACKGROUND: Human basic fibroblast growth factor (hBFGF) is a heparin-binding growth factor and stimulates the proliferation of a wide variety of cells and tissues causing survival properties and its stability and biological activity improvements have received much attention. MATERIALS AND METHODS: In the present work, hBFGF produced by engineered Escherichia coli and purified by cation exchange and heparin affinity chromatography, was PEGylated under appropriate condition employing 10 kD polyethylene glycol. The PEGylated form was separated by size exclusion chromatography. Structural, biological activity, and stability evaluations were performed using Fourier transform infrared (FITR) spectroscopy, 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) assay and effect denaturing agent, respectively. RESULTS: FITR spectroscopy revealed that both PEGylated and native forms had the same structures. MTT assay showed that PEGyalated form had a 30% reduced biological activity. Fluorescence spectrophotometry indicated that the PEGylated form denatured at higher concentrations of guanidine HCl (1.2 M) compared with native, which denatured at 0.8 M guanidine HCl. CONCLUSIONS: PEGylation of hBFGF makes it more stable against denaturing agent but reduces its bioactivity up to 30%.

Effects of mechanical and chemical stimuli on differentiation of human adipose-derived stem cells into endothelial cells.

It has been hypothesized that application of the micromechanical environment that target cells experience in vivo enhances functionality of differentiated cells. Vascular endothelial cells, functioning at the interface of the blood-vessel wall, are vital to the performance of the cardiovascular system. They are subject to shear and tensile stresses induced by blood flow and pressure, respectively. This study investigated effects of shear/tensile stresses on endothelial differentiation of adipose-derived mesenchymal stem cells (ASCs) utilizing a custom-made bioreactor capable of applying both shear and tensile stresses. The loading values of 10% cyclic stretch, 0-2.5 dyn/cm² cyclic shear stress, and combined loadings were used. To examine the extent of mechanical and chemical stimuli in acquisition of endothelial characteristics by ASCs, the expression of three major endothelial genes were quantified when ASCs were treated by three loading regimes and endothelial growth factor for three different durations (1, 2, and 7 days). In general, cyclic stretch decreased expression of FLK-1 and vWF, while cyclic shear elevated expression levels. The combined loading regime had minor effects on the expression of the two markers. All types of loadings significantly enhanced the expression level of VE-cadherin with the most prominent increase by combined loading. It was concluded that applying different loading regimes assists in adjusting the expression level of endothelial markers to achieve functional endothelial cells for cardiovascular engineering.

Cytotoxic effects of human calprotectin on gastric cancer cell line is attenuated by etoposide.

AIM: In this paper effect of combinational usage of calprotectin and etoposide on AGS cell line is studied. BACKGROUND: Application of combined toxic agents such as etoposide and cicplatin are commonly used for chemotherapy purposes. As a matter of fact, calprotectin and etoposide were both applied on human gastric adenocarcinoma cell line (AGS) as antitumor agents. Both calprotectin and etoposide are topo II inhibitor. Etoposide is a lipophilic agent that can easily transport from membrane while calprotectin active intracellular pathway, probably by membrane surface receptor. PATIENTS AND METHODS: Calprotectin was purified from human neutrophil by chromatography methods. The human gastric adenocarcinoma cell line was exposed to different concentrations and combinations of calprotectin and etoposide. MTT assay was applied for evaluation of cytotoxicity assay. RESULTS: Viability of AGS cell line was reduced in high dosages of calprotectin and etposide. In fact, overnight incubation of these two agents together has been shown less effective than individual usage. CONCLUSION: The result indicates that, the combination of both calprotectin and etoposide is considerably less cytotoxic on gastric cancer cells (AGS) than applying individually.

High-level expression of functional recombinant human coagulation factor VII in insect cells.

Recombinant coagulation factor VII (FVII) is used as a potential therapeutic intervention in hemophilia patients who produce antibodies against the coagulation factors. Mammalian cell lines provide low levels of expression, however, the Spodoptera frugiperda Sf9 cell line and baculovirus expression system are powerful systems for high-level expression of recombinant proteins, but due to the lack of endogenous vitamin K-dependent carboxylase, expression of functional FVII using this system is impossible. In the present study, we report a simple but versatile method to overcome the defect for high-level expression of the functional recombinant coagulation FVII in Sf9 cells. This method involves simultaneous expression of both human gamma-carboxylase (hGC) and human FVII genes in the host. It may be possible to express other vitamin K-dependent coagulation factors using this method in the future.

Immunochemical characterization of alkaline phosphatase from the fluid of sterile and fertile Echinococcus granulosus cysts.

Alkaline phosphatase (ALP) from hydatid cyst fluid (HCF) was purified and characterized for comparison between fertile and sterile HCF. Samples were obtained from slaughtered sheep and then sterile and fertile cysts were separated. ALP was purified from aspirated cyst fluid and biochemical parameters were determined. Sera from patients with hydatid disease (15 samples) and patients with other parasitic diseases including fascioliasis (2 samples), taeniasis ( Taenia saginata, 5 samples) and also sera from uninfected controls (15 samples), were collected and used in immunoblotting experiments with ALP from sterile and fertile HCF as antigen. Our results showed that ALP activity in fertile HCF [10.75+/-3.78 (SD) U/ml) was significantly more than in sterile HCF (6.25+/-2.43 U/ml). There were also some differences between the kinetic parameters and biochemical characteristics of ALP in fertile and sterile HCF. Immunoreactive bands were clearly observed when sera from hydatid infected patients were tested with ALP from fertile HCF as the antigen. However, this method revealed no cross-reaction between purified ALP from sterile HCF and sheep liver tissue. These findings suggest that there is some variation in the immunochemical characteristics of ALP from fertile and sterile HCF.