Osteoblast Differentiation on Collagen Scaffold with Immobilized Alkaline Phosphatase.

BACKGROUND: In tissue engineering, scaffold characteristics play an important role in the biological interactions between cells and the scaffold. Cell adhesion, proliferation, and activation depend on material properties used for the fabrication of scaffolds. OBJECTIVE: In the present investigation, we used collagen with proper characteristics including mechanically stability, biodegradability and low antigenicity. Optimization of the scaffold was done by immobilization of alkaline phosphatase on the collagen surface via cross-linking method, because this enzyme is one of the most important markers of osteoblast, which increases inorganic phosphate concentration and promote mineralization of bone formation. METHODS: Alkaline phosphatase was immobilized on a collagen surface by 1-ethyl-3-(dimethylaminopropyl) carbodiimide hydrochloride, as a reagent. Then, rat mesenchymal stem cells were cultured in osteogenic medium in control and treated groups. The osteogenesis-related genes were compared between treatments (differentiated cells with immobilized alkaline phosphatase/collagen scaffold) and control groups (differentiated cells on collagen surface without alkaline phosphatase) on days 3 and 7 by quantitative real-time PCR (QIAGEN software). RESULTS: Several genes, including alkaline phosphatase, collagen type I and osteocalcine associated with calcium binding and mineralization, showed upregulation in expression during the first 3 days, whereas tumor necrosis factor-α, acting as an inhibitor of differentiation, was down-regulated during osteogenesis. CONCLUSION: Collagen scaffold with immobilized alkaline phosphatase can be utilized as a good candidate for enhancing the differentiation of osteoblasts from mesenchymal stem cells.

Conservation and diversity in the structure of translation initiation factor EIF3 from humans and yeast.

Initiation factor eIF3 plays a central role in the initiation pathway, influencing ribosome association, ternary complex binding to 40S subunits, and mRNA binding, in part through an interaction with eIF4F. We are attempting to clone and sequence DNAs encoding the subunits of this complex factor. Mammalian eIF3 comprises 10 subunits; full-length human cDNAs have been cloned for eight of these, and partial clones are in hand for the remaining two. Yeast eIF3 comprises at least seven subunits, with six of the seven genes identified and sequenced. Comparison of eIF3 subunit sequences between human and yeast reveals an unexpectedly large diversity of structure. Surprisingly, comparisons with other sequences in the data base suggest that some of the eIF3 subunits may have functions apart from the eIF3 complex. Work is in progress to use the cloned DNAs as tools for elucidating the structure of eIF3 and its interactions with other initiation factors.

The effect of neem (Azadirachta indica) extract and dietary selenium on distribution of selenium in hepatocarcinogenesis induced rat.

Neem, Azadirachta indica, is a plant from the family Meliaceae, known as "Pokok Semambu" in Malay community. It has been extensively used in India as traditional Ayurvedic and folklore minedicine for the treatment of various diseases. This study aimed to determine the distribution of selenium in the liver of rats during hepatocarcinogenesis when neem aqueous extract and dietary selenium was supplemented.

Effects of Soy on Body Composition: A 12-Week Randomized Controlled Trial among Iranian Elderly Women with Metabolic Syndrome.

BACKGROUND: To examine the effects of soy [in the form of textured soy protein (TSP) and soy-nut] on body composition in elderly women with metabolic syndrome (MetS). METHODS: A 12-week randomized clinical trial was conducted on 75 women between 60-70 years of age with MetS in rural health clinics around Babol, Iran in 2009. The participants were randomly assigned to one of the three groups of soy-nut (35g/d), TSP (35g/d) and control. Body fat, lean mass and anthropometric indicators were measured before and after intervention, too. RESULTS: Participants were classified as overweight and showing android fat distribution. After 12 weeks of intervention, both soy-nut and TSP groups showed an increase of non-significant in lean mass (0.9 and 0.7 kg), hip circumference (0.45 and 0.28 cm), triceps skinfold (TSF) thickness (0.87 and 0.67mm) and reduction in BMI (-0.15 and -0.33), waist circumference (-0.83 and -1.2) and body fat (-1.5% and -1.7%). Significant increase in the mean change of TSF and lean mass was observed in the users of soy-nut compared to the control group (P<0.01, P<0.05). CONCLUSION: 12-week intervention of soy had a mild favorable effect on body composition in elderly women with MetS.

Physical activity, nutrition, and dyslipidemia in middle-aged women.

BACKGROUND: Cardiovascular disease is a major cause of death throughout the world. The aim of this study was to assess the prevalence of overweight/obesity, central obesity, hypertension and dyslipidemia, as well as dietary factors contributing to the development of dyslipidemia among middle-aged women. METHODS: The research design of the present study was a population-based cross-sectional study; anthropometric measures and blood chemistry were obtained. Physical activity was measured using the original International Physical Activity Questionnaires Long Form while food frequency questionnaire (FFQ) was used in assessing individual's habitual intake. Overall, 809 women, 30-50 years of age from fourteen active urban Primary Healthcare Centers (PHC) in Babol City, northern Iran, were obtained from 1,905 households across operational areas of 14 PHC using systematic random sampling method. RESULTS: The prevalence rates of women classified as overweight/obese, with central obesity, hypertension and dyslipidemia were 82.8%, 75.5%, 14.6% and 63.4%, respectively. Total physical activity did not correlate with cholesterol ratio. Soybean protein was inversely associated with cholesterol ratio (rho=-0.18, P≤ 0.001). The adjusted OR for dyslipidemia in women with moderate protein intake was significantly higher than in women with high and low intake (OR=2.31; 95% CI= 1.61, 3.30). No significant associations were found between dyslipidemia and carbohydrate, fat intake or physical activity. CONCLUSION: This study showed very high prevalence of cardiovascular disease risk factors among Iranian middle-aged women. A more detailed study is suggested to develop definitively recommendations for the primary and secondary prevention of cardiovascular disease for the Iranian population.

Determination of daidzein and genistein in soy milk in Iran by using HPLC analysis method.

The HPLC system separated completely isofavonoids such as daidzein (15.2 min) and genistein (17.3 min). Initially, the concentrations of major isoflavone Genistein and Daidzein in the tested soy milk were determined. Commercial soy milk samples were analyzed for isoflavones and two major isoflavones detected: genistein 25.86 (mg L(-1)) +/- 0.66 SD and daidzein 8.25 (mg L(-1)) +/- 1.13 SD. Concentrations of genistein in soy milk were higher than daidzein. The results obtained in this study can serve as a basis for estimating amount of soy milk can be consumed by people as related to its main isoflavone content.

Characterization of the p33 subunit of eukaryotic translation initiation factor-3 from Saccharomyces cerevisiae.

Eukaryotic translation initiation factor-3 (eIF3) is a large multisubunit complex that binds to the 40 S ribosomal subunit and promotes the binding of methionyl-tRNAi and mRNA. The molecular mechanism by which eIF3 exerts these functions is incompletely understood. We report here the cloning and characterization of TIF35, the Saccharomyces cerevisiae gene encoding the p33 subunit of eIF3. p33 is an essential protein of 30,501 Da that is required in vivo for initiation of protein synthesis. Glucose repression of TIF35 expressed from a GAL1 promoter results in depletion of both the p33 and p39 subunits. Expression of histidine-tagged p33 in yeast in combination with Ni2+ affinity chromatography allows the isolation of a complex containing the p135, p110, p90, p39, and p33 subunits of eIF3. The p33 subunit binds both mRNA and rRNA fragments due to an RNA recognition motif near its C terminus. Deletion of the C-terminal 71 amino acid residues causes loss of RNA binding, but expression of the truncated form as the sole source of p33 nevertheless supports the slow growth of yeast. These results indicate that the p33 subunit of eIF3 plays an important role in the initiation phase of protein synthesis and that its RNA-binding domain is required for optimal activity.

Identification of partners of TIF34, a component of the yeast eIF3 complex, required for cell proliferation and translation initiation.

Eukaryotic initiation factor-3 (eIF3) in the yeast Saccharomyces cerevisiae plays a central role in initiation of translation. The eIF3 complex contains at least eight different proteins, but, as yet, little is known about the function of the individual proteins. In this study we have characterized the role of TIF34 (eIF3-p39), a recently identified WD-40 domain-containing protein of 39 kDa, in the eIF3 complex. Using temperature-sensitive mutants of TIF34 we show that this protein is required for cell cycle progression and for mating and plays an essential role in initiation of protein synthesis. By two-hybrid screening we have identified two partners that directly associate with TIF34: PRT1, a previously characterized eIF3 subunit, and a novel protein of 33 kDa (eIF3-p33) which is part of the eIF3 complex and has an RNA binding domain. TIF34 and p33 interact with each other and overexpression of p33 complements the growth defect of a tif34-ts mutant. Our results provide support for both physical and functional interactions between three subunits, TIF34, PRT1 and p33, in the eIF3 complex.

A 110-kilodalton subunit of translation initiation factor eIF3 and an associated 135-kilodalton protein are encoded by the Saccharomyces cerevisiae TIF32 and TIF31 genes.

Translation initiation factor eIF3 is a multisubunit protein complex required for initiation of protein biosynthesis in eukaryotic cells. The complex promotes ribosome dissociation, the binding of the initiator methionyl-tRNA to the 40 S ribosomal subunit, and mRNA recruitment to the ribosome. In the yeast Saccharomyces cerevisiae eIF3 comprises up to 8 subunits. Using partial peptide sequences generated from proteins in purified eIF3, we cloned the TIF31 and TIF32 genes encoding 135- (p135) and 110-kDa (p110) proteins. Deletion/disruption of TIF31 results in no change in growth rate, whereas deletion of TIF32 is lethal. Depletion of p110 causes a severe reduction in cell growth and protein synthesis rates as well as runoff of ribosomes from polysomes, indicative of inhibition of the initiation phase. In addition, p110 depletion leads to p90 co-depletion, whereas other eIF3 subunit levels are not affected. Immunoprecipitation or nickel affinity chromatography from strains expressing (His)6-tagged p110 or p33 results in the co-purification of the well characterized p39 and p90 subunits of eIF3 as well as p110 and p33. This establishes p110 as an authentic subunit of eIF3. In similar experiments, p135 and other eIF3 subunits sometimes, but not always, co-purify, making assignment of p135 as an eIF3 subunit uncertain. Far Western blotting and two-hybrid analyses detect a direct interaction of p110 with p90, p135 with p33, and p33 with eIF4B. Our results, together with those from other laboratories, complete the cloning and characterization of all of the yeast eIF3 subunits.