Cell attachment and proliferation of human adipose-derived stem cells on PLGA/Chitosan Electrospun Nano-Biocomposite

Objective: In this study, nano-biocomposite composed of poly [lactide-co-glycolide] [PLGA] and chitosan [CS] were electrospun through a single nozzle by dispersing the CS nano-powders in PLGA solution. The cellular behavior of human adipose derived stem cells [h-ADSCs] on random and aligned scaffolds was then evaluated

Materials and Methods: In this experimental study, the PLGA/CS scaffolds were prepared at the different ratios of 90/10, 80/20, and 70/30 [w/w]%. Morphology, cell adhesion and proliferation rate of h-ADSCs on the scaffolds were assessed using scanning electron microscope [SEM], 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide [MTT] assay and trypan blue staining respectively

Results: H-ADSCs seeded on the matrices indicated that the PLGA/CS composite matrix with aligned nanofibres and higher content of CS nano-powders gave significantly better performance than others in terms of cell adhesion and proliferation rate [P<0.05]

Conclusion: We found that CS enhanced cell adhesion and proliferation rate, and aligned nanofibers guided cell growth along the longitudinal axis of the nanofibers, which would provide a beneficial approach for tissue engineering

Comparative study of microtubule-associated protein-2 and glial fibrillary acidic proteins during neural induction of human bone marrow mesenchymal stem cells and adipose-derived stem cells

In recent years, adipose tissue, due to the stem cells contained within, has found a new special place in laboratory and clinical applications. These adipose-derived stem cells [ADSCs] have the same characteristics of bone marrow mesenchymal stem cells [BMSCs]. Although bone marrow [BM] is not easily accessible and its procurements may be painful, most patients possess excess fat which can be obtained by less invasive methods; this makes adipose tissue ubiquitous, available and an ideal large-scale source for research on clinical applications. BMSCs and ADSCs were harvested from three healthy human and were characterized using flow-cytometry. After they were treated for neurosphere formation using basic fibroblast growth factor, epidermal growth factor, B27; terminal differentiation was performed. In this study, we used immunocytochemistry, real time-polymerase chain reaction and western blotting techniques for detection and comparison of Nestin, microtubule-associated protein-2 [MAP-2] and glial fibrillary acidic protein [GFAP] markers in human ADSCs and BMSCs. Under appropriate conditions ADSCs can differentiate into neuron-like cells and express neural markers the same as BMSCs, also the expression of GFAP marker in differentiated cells derived from ADSCs was significantly lower than the cells derived from BMSCs [P < 0.05]. While the expression of MAP-2 marker in both groups was the same. However, due to its advantages and according to our results based on the expression levels of GFAP and MAP-2, adipose tissue rather than BM could represent a more appropriate stem cell source for investigating the application of these cells in understanding the pathophysiology and in treatment of neurodegenerative disorders

Preparation and cytotoxic evaluation of magnetite [Fe[3]O[4]] nanoparticles on breast cancer cells and its combinatory effects with doxorubicin used in Hyperthermia

Magnetic nanoparticles in a variable magnetic field are able to produce heat. This heat [42-45[degree sign]C] has more selective effect on fast dividing cancer cells than normal tissues. In this work magnetite nanoparticles have been prepared via coprecipitation and phase identification was performed by powder x-ray diffraction [XRD]. Magnetic parameters of the prepared nanoparticles were measured by a Vibrating Sample Magnetometer [VSM]. A sensitive thermometer has been used to measure the increase of temperature in the presence of an alternating magnetic field. To evaluate the cytotoxicity of nanoparticles, the suspended magnetite nanoparticles in liquid paraffin, doxorubicin and a mixture of both were added to the MDA-MB-468 cells in separate 15 ml tubes and left either in the RT or in the magnetic field for 30 min. Cell survival was measured by trypan blue exclusion assay and flow cytometer. Particle size distribution of the nanoparticles was homogeneous with a mean particles size of 10 nm. A 15[degree sign]C temperature increase was achieved in presence of an AC magnetic field after 15 min irradiation. Biological results showed that magnetite nanoparticles alone were not cytotoxic at RT, while in the alternative magnetic filed more than 50% of cells were dead. Doxorubicin alone was not cytotoxic during 30 min, but in combination with magnetite more than 80% of the cells were killed. It could be concluded that doxorubicin and magnetite nanoparticles in an AC magnetic field had combinatory effects against cells

[Production of recombinant human growth hormone by eukaryotic CHO cell and measurement of its biological activity by gene reporter assay]

Cultivated mammalian cells, because of their capacity for proper protein folding, assembly and post-translational modification, have become the dominant system for production of recombinant proteins in clinical application. Therefore, the quality and efficacy of protein can be superior when expressed in mammalian cells compared to other hosts such as bacteria. Gene reporter systems have contributed greatly to the study of eukaryotic gene expression and regulation. Although reporter genes have played a significant role in numerous applications, both in vitro and in vivo, they are most frequently used as indicators of transcriptional activity in cells. Luciferase-reporter assays are widely used to monitor the cellular events related to transduction and gene expression regulated by specific cascades, such as PRL/Jak2/Stat 5 pathway. In this study, recombinant human growth hormone [rhGH] was produced in eukaryotic Chinese hamster ovary [CHO] cell and production and concentration of rhGH verified by ELISA and western blotting. Then, the biological activity of rhGH was assessed by a gene reporter assay system [containing LHRE, TK promoter and Luc gene], using HEK 293 cells transfected with GH receptor and response element for STAT-5 measuring luciferase activity on a Berthold luminometer. The date showed that rhGH could be produced by eukaryotic host in good quantities as assessed by ELISA and western blotting. The results of gene reporter assay showed that rhGH produced by CHO cells is able to induce GH intracellular cell signaling. The rhGH produced by CHO cells showed higher bioactivity when compared to commercial GH. rhGH could be produced in mammalian cell lines at high levels with higher bioactivity. Gene reporter assay is a sensitive, quantitative, rapid, easy, reproducible and safe system for assessment of bioactivity of recombinant proteins such as rhGH

Antibodies to Interferon beta in Patients with Multiple Sclerosis Receiving CinnoVex, Rebif, and Betaferon

Treatment with interferon beta (IFN-beta) induces the production of binding antibodies (BAbs) and neutralizing antibodies (NAbs) in patients with multiple sclerosis (MS). NAbs against IFN-beta are associated with a loss of IFN-beta bioactivity and decreased clinical efficacy of the drug. The objective of this study was to evaluate the incidence and the prevalence of binding antibodies (BAbs) and neutralizing antibodies (NAbs) to IFN-beta in MS patients receiving CinnoVex, Rebif, or Betaferon. The presence of BAbs was studied in serum samples from 124 MS patients using one of these IFN-beta medications by ELISA. The NAbs against IFN-beta were measured in BAb-positive MS patients receiving IFN-beta using an MxA gene expression assay (real-time RT-PCR). Of the 124 patients, 36 (29.03%) had BAbs after at least 12 months of IFN-beta treatment. The proportion of BAb+ was 38.1% for Betaferon, 21.9% for Rebif, and 26.8% for CinnoVex. Five BAb-positive MS patients were lost to follow-up; thus 31 BAb-positive MS patients were studied for NAbs. NAbs were present in 25 (80.6%) of BAb-positive MS patients receiving IFN-beta. In conclusion, the three IFN-beta preparations have different degrees of immunogenicity.

Characterization of an interesting novel mutant strain of commercial Saccharomyces cerevisiae

The yeast strains that are resistant to high concentration of ethanol have biotechnological benefits and are suitable models for physiology and molecular genetics research fields. A novel ethanol-tolerant mutant strain, mut1, derived from the commercial Saccharomyces cerevisiae showed higher ethanol production, and also demonstrated resistance to ethanol but not to other alcohols, such as methanol, 2-propanol, and 1-butanol. To characterize mut1, the strain's resistance to other organic compounds and osmotic and cell wall stresses were examined. The growth of the mut1 strain in the presence of ethyl n-caproate and 3-methyl butyl acetate, which were metabolic derivatives of ethanol, was found to be less than the wild type. On the other hand, the growth of the mut1 strain in the presence of 50% [w/v] sucrose and 1M NaCl was similar to that of the wild type. The sensitivity to cell wall digestive enzyme, zymolyase, was also similar in both wild and mut1 strains. Finally, the mut1 strain showed resistance to homocysteine and serine but was sensitive to methionine. These results suggest that the ethanol resistance of the mut1 strain may be more related to the ethanol metabolic and signalling pathways rather than the enhanced stress resistances relating to the membrane or cell wall compositions

[Prevalence of nano structure S-layer and beta-lactamase in Bacillus cereus strains]

S-layer is the outer protein layer in the most archaea and bacteria. S-layer protects bacteria against phagocytosis and prohibits entry of some biomolecules and some antibiotics and adhesion to matrix proteins. S-layer is a virulence agent in bacteria, beta-lactamase can inactive [beta-lactame antibiotics. According to role of beta-lactame antibiotics in the treatment of infectious diseases with Bacillus spp., increasing frequency of S-layer and beta-lactamase producer Bacillus cereus strains in hospitals lead to increase antibiotic resistant nosocomial infections. In this basic study, 274 samples were evaluated with laboratory methods in Alzahra hospital and Isfahan University between 2004 and 2006. Bacterial identification was performed with microbiological methods, such as staining, chemical test, use of differential and selective Bacillus cereus selective agar media. Bacteria cultured in TSA for 16h, and then separated surface proteins, and finally electrophoresis was performed. S-layer in Bacillus cereus has 97KD molecular weight. Production of beta-lactamase was evaluated by acidometric method. Of 247 isolated bacteria, frequency of Bacillus cereus strains, S-layer and beta-lactamase in S-layer producer Bacillus cereus strains were 9.49%, 46.20% and 100%, respectively. Results showed high prevalence of nano structure S-layer and beta-lactamase producer Bacillus cereus strains in hospital. We recommend controlling bacterial population in crowded places and health and therapeutic centers to decrease producing antibiotic resistance bacteria