فهرست مطالب

Biotechnology - Volume:14 Issue: 3, Summer 2016

Iranian Journal of Biotechnology
Volume:14 Issue: 3, Summer 2016

  • تاریخ انتشار: 1395/07/24
  • تعداد عناوین: 9
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  • Shakeel Ahmed Ansari*, Rukhsana Satar, Mohammad Alam Jafri, Mahmood Rasool, Waseem Ahmad, Syed Kashif Zaidi Pages 70-81
    Context: The use of nanotechnology in medicine and more specifically drug delivery is set to spread rapidly. Currently many substances are under investigation for drug delivery and more specifically for cancer therapy.
    Evidence Acquisition: Nanodiamonds (NDs) have contributed significantly in the development of highly efficient and successful drug delivery systems, and in stem cell therapy. Drug delivery through NDs is an intricate and complex process that deserves special attention to unravel underlying molecular mechanisms in order to overcome certain bottlenecks associated with it. It has already been established that NDs based drug delivery systems have excellent biocompatibility, non-toxicity, photostability and facile surface functionalization properties.
    Results
    There is mounting evidence that suggests that such conjugated delivery systems well retain the properties of nanoparticles like small size, large surface area to volume ratio that provide greater biocatalytic activity to the attached drug in terms of selectivity, loading and stability.
    Conclusions
    NDs based drug delivery systems may form the basis for the development of effective novel drug delivery vehicles with salient features that may facilitate their utility in fluorescence imaging, target specificity and sustained-release.
    Keywords: Biomedical applications, Drug delivery, Hydrogels, Nanodiamonds, Surface functionalization
  • Arastoo Badoei, Dalfard*, Narjes Ramezani, Pour, Zahra Karami Pages 82-93
    Background
    The conversion of nitriles into amides or carboxylic acids by nitrilase has taken its application into consideration, as the scope of its applications has recently been extended.
    Objectives
    In this study, P. aeruginosa RZ44 was isolated from sewage in the Kerman which has Nitrile-degradation activity. In order to improve the nitrilase production, several optimization were done on environmental condition. Nitrilase activity was characterized against different pHs, temperatures, ions, and substrates.
    Materials And Methods
    Enzyme activity was evaluated by determining the production of ammonia following to the modification of the phenol/hypochlorite method. Different factors that affect production of the enzyme by P. aeruginosa RZ44 were optimized and evaluated in the culture mediums.
    Results
    The results showed that degradation of the acetonitrile by P. aeruginosa RZ44 increased the pH of the growth medium from the initial pH 7.0 to 9.37. Optimizing the medium for P. aeruginosa RZ44, it was found that glucose and starch (5 g.L-1) have strongly supported nitrilase production, compared to the control. As well, urea (5 g.L-1) and yeast extract (15 g.L-1) have favored an increased biomass and nitrilase production, as the nitrogen sources. These results show that nitrilase production increases in the pH range 5.0 to 7.0 and then start decreasing. Addition of the Mg2, Fe2 and Na has supported the biomass and nitrilase production. Co2, Mn2 and Cu2 were confirmed to inhibit cell growth and enzyme production. Enzyme characterization results show that, P. aeruginosa RZ44 nitrilase exhibits comparatively high activity and stability at pH 7.0 and 40°C. Nitrilase was completely inhibited by CoCl2 and CaCl2, whereas, the inhibition in the presence of MnSO4 and CuSO4 was about 60%. Time course analysis of the nitrile conversion by the resting P. aeruginosa RZ44 cells showed that nitrile substrates (i.e. acetonitrile) was hydrolyzed within 8 h.
    Conclusions
    these results indicate that P. aeruginosa RZ44 has the potential to be applied in the biotransformation of nitrile compounds.
    Keywords: Nitrilase, Nitrile, Nitrile, degrading bacteria, Production, Pseudomonas aeruginosa, Sewage
  • Kaizar Hossain, Shlrene Quaik, Norli Ismail*, Mohd Rafatullah, Maruthi Avasan, Rameeja Shaik Pages 94-102
    Background
    Application of membrane technology to wastewater treatment has expanded over the last decades due to increasingly stringent legislation, greater opportunities for water reuse/recycling processes and continuing advancement in membrane technology.
    Objectives
    In the present study, a bench-scale submerged microfiltration membrane bioreactor (MBR) was used to assess the treatment of textile wastewater.
    Materials And Methods
    The decolorization capacity of white-rot fungus coriolus versicolor was confirmed through agar plate and liquid batch studies. The temperature and pH of the reactor were controlled at 29±1°C and 4.5±2, respectively. The bioreactor was operated with an average flux of 0.05 m.d-1 (HRT=15hrs) for a month.
    Results
    Extensive growth of fungi and their attachment to the membrane led to its fouling and associated increase of the transmembrane pressure requiring a periodic withdrawal of sludge and membrane cleaning. However, stable decoloration activity (approx. 98%), BOD (40-50%), COD (50-67%) and total organic carbon (TOC) removal (>95%) was achieved using the entire system (fungi membrane), while the contribution of the fungi culture alone for TOC removal, as indicated by the quality of the reactor supernatant, was 35-50% and 70%, respectively.
    Conclusions
    The treated wastewater quality satisfied the requirement of water quality for dyeing and finishing process excluding light coloration. Therefore, textile wastewater reclamation and reuse is a promising alternative, which can both conserve or supplement the available water resource and reduce or eliminate the environmental pollution.
    Keywords: Bioremediation, Membrane bioreactor, Textile wastewater, White, rot fungi
  • Farajollah Ahmadi*, Abbas Tanhaeian, Maziar Habibi, Pirkoohi Pages 103-108
    Background
    Biosynthesis of nanoparticles using microorganisms, enzymes, and plant extracts is regarded as an alternative to chemical methods. Microalgae appear to be an efficient biological platform for nanoparticle synthesis as they grow rapidly and produce large biomass at lower cost.
    Objectives
    The possibility of silver nanoparticles biosynthesisby freshwater green microalgae, Chlamydomonas reinhardtii, was evaluated. Furthermore, antibacterial properties of the synthesized nanoparticles were investigated via analysis of growth and toxin production of Listeria monocytogenes.
    Materials And Methods
    Silver nanoparticles were synthesized by incubating 47.5 mL of fresh C. reinhardtii culture with 2.5 mL of 200 mM AgNO3 solution for 48 h. Characterization of the synthesized nano particles was performed by Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Energy Dispersive Spectrometry (EDS) and X-ray diffraction analysis (XRD). Concentration of biosynthesized silver nanoparticles was measured by high resolution ICP-OES spectrometer. Inhibitory effect of silver nanoparticles on L. monocytogenes growth was measured. Further, the expression of listeriolysin O was investigated by serial microdilution method and Real-Time PCR assay.
    Results
    Spherical silver nanoparticles with average size of about 10 nm were formed. The particles had inhibitory effects on bacterial growth and antagonist activity on the expression of listeriolysin O.
    Conclusions
    C. reinhardtii has the potential to be used as an effective platform for production of silver and other nanoparticles. Silver nanoparticles had potent antibacterial properties.
    Keywords: Biosynthesis, Chlamydomonas reinhardtii, Listeriosis
  • Khadijeh Karbalaie, Sadeq Vallian*, Liana Lachinani, Somayeh Tanhaei, Hossein Baharvand, Mohammad Hossein Nasr, Esfahani Pages 109-116
    Background
    Promyelocytic leukemia protein (PML) is a tumor suppressor protein that is involved in myeloid cell differentiation in response to retinoic acid (RA). In addition, RA acts as a natural morphogen in neural development.
    Objectives
    This study aimed to examine PML gene expression in different stages of in vitro neural differentiation of NT2 cells, and to investigate the possible role of PML in pluripotency and/or neural development.
    Materials And Methods
    RA was used as a neural inducer for in vitro neural differentiation of NT2 cells. During this process PML mRNA and protein levels were assessed by quantitative real time RT-PCR (QRT-PCR) and Immunoblotting, respectively. Furthermore bisulfite sequencing PCR (BSP) was used to assess PML promoter methylation in NT2 cells and NT2 derived neuronal precursor cells (NT2.NPCs).
    Results
    QRT-PCR results showed that, PML had maximum expression with significant differences in NT2 derived neuronal precursor cells relative to NT2 cells and NT2 derived neural cells (NT2.NCs). Numerous isoforms of PML with different intensities appeared in immunoblots of pluripotent NT2 cells, NT2.NPCs, and NT2.NCs. Furthermore, the methylation of the PML promoter in NT2.NCs was 2.6 percent lower than NT2 cell.
    Conclusions
    The observed differences in PML expression in different cellular stages possibly could be attributed to the fact that PML in each developmental state might be involved in different cell signaling machinery and different functions. The appearance of different PML isoforms with more intensity in neural progenitor cells; may suggest apossible role for this protein in neural development.
    Keywords: Pluripotent stem cells, Promyelocytic Leukemia, Retinoic acid
  • Maliheh Bahadori, Javad Baharara*, Elaheh Amini Pages 117-124
    Background
    The SALL4/Sall4 is constitutively expressed in human and mice. SALL4 mRNA could be used as a marker for the diagnosis of different types of cancers. On the other hand, chrysin has diverse biological properties.
    Objectives
    In the present study, the effect of the chrysin was investigated on the CT26 colon cancer in vitro and in vivo. Furthermore, the expression levels of the stem cell markers; sall4 and Bax was analyzed, as well.
    Materials And Methods
    The cytotoxic effects and the type of cell death induced by chrysin were evaluated using a number of biological assays. The apoptotic pathway was examined by caspase-3and caspase-9 assay. The in vivo antitumor efficacy of chrysin on transplanted CT26 tumor cells in BALB/c mice was investigated. In addition, mRNA expression of sall4, Bax was analyzed with RT-PCR.
    Results
    MTT assay and morphological characteristics showed that chrysin exerted a cytotoxic effect on CT26 cells in a dose dependent manner with IC50= 80 mg.mL-1. The biological assays have indicated that chrysin administrated cytotoxicity on colon cancer cells through recruitment of the apoptosis. Caspase-3 and caspase-9 colorimetric assays, in addition to Bax expression analysis, have indicated the involvement of intrinsic apoptotic pathway in the cytotoxic effect of the chrysin. The in vivo assay revealed a remarkable reduction of the colon tumor volume in treated mice (8, 10 mg.kg -1) as compared to the untreated mice. RT-PCR elucidated that chrysin attenuated tumor volume through down regulation of the sall4 and up-regulation of the Bax.
    Conclusions
    It was demonstrated that chrysin accomplishes anti-cancer effect on colon cancer cells via induction of the apoptosis and attenuation of the sall4 the expression. These findings introduce chrysin as an efficient apoptosis based therapeutic agent against colon cancer.
    Keywords: Apoptosis, Chrysin, Colon cancer, In vivo, Sall4
  • Shahryar Khoshtinat Nikkhoi, Ruhollah Dorostkar, Saeed Ranjbar, Hedieh Heydarzadeh, Mahdi Tat, Majdedin Ghalavand, Alireza Farasat, Mohammad Sadegh Hashemzadeh* Pages 125-131
    Background
    Puma is a highly robust pro-apoptotic protein. The protein becomes activated by p53 ensuing beyond-repair DNA damage. Downregulation of SIRT 1, by miR-128, elevates activated p53 that foment Puma indirectly.
    Objectives
    In the present study, we used two-expression Adeno-Associated Virus (AAV) system for co-expression of miR-128 and Puma in order to evaluate apoptotic response; both in the tumor and normal cells, respectively.
    Materials And Methods
    Three recombinant AAVs constructs were generated. The First rAAV bearing Puma under the control of hTERT (p-AAV), the second construct designed such that to carry miR-128 downstream of CMV (mi-AAV), and the last construct comprises of the both CMV-miR-128 and hTERT- Puma. Real-Time PCR and western blotting were used to evaluate expression levels of the transduced genes.
    Results
    MTT assay and DAPI staining shown suicidal effect of each recombinant AAV vectors. p-AAV cytotoxicity was recorded for 62% of the tumor cells, while for normal cells it was only 20% cytotoxic. The second construct, mi-AAV, was not as potent and selective as p-AAV. This construct was shown to be 27% and 16% cytotoxic for BT-474 and HEK-293 cells, respectively. Co-expression of Puma and miR-128 (p-mi-AAV) was accomplished with a selective cytotoxicity toward BT-474. This construct was 85% toxic for tumor cells, although it was only 25% toxic for the normal cell line (HEK-293).
    Conclusions
    In this study, we have shown that not only Puma is able to instigate apoptotic response but also its co-expression along with miR-128 could significantly enhance apoptosis in a synergistic manner.
    Keywords: AAV, Adeno, Associated Virus, Gene therapy, Puma, Suicide gene
  • Hamida Bouridane, Mohamed Sifour, Tayeb Idoui *, Lejeune Annick, Philip Thonard Pages 132-141
    Background
    For biotechnological application, selected lactic acid bacteria strains belonging to the genera Lactobacillus (Lb) are proposed as an alternative to the antibiotics for the prevention and treatment of urogenital tract infections.
    Objectives
    Isolating and selecting vaginal lactobacilli strains for probiotic use based on their technological and probiotic aptitudes.
    Materials And Methods
    The vaginal isolates were examined for their essential characteristics as the potential probiotic such as low pH tolerance, bile-salt and simulated human intestinal fluid (SIF) resistance, adhesion to the vaginal epithelial cells (VECs), aggregation and coaggregation, surface hydrophobicity, antimicrobial activity, acid production, antibiotic resistance, and resistance to spermicides. The best strain was identified by PCR.
    Results
    From 70 lactobacilli isolates and according to the 16 rDNA sequences, isolates B6 and B10 showed the closest homology (99%) to the Lb. gasseri and Lb. plantarum respectively. They produced hydrogen peroxide (H2O2), tolerant to acid, bile, simulated human intestinal fluid, present a strong adhesion, highest percentages of aggregation, and antibacterial activity. These strains are resistant to the spermicide and actively acidify the growth medium.
    Conclusions
    Strains Lb. plantarum B10 and Lb. gasseri B6 have a strong potential probiotic confirming their value as a tool for prevention against urinary and vaginal infections.
    Keywords: Adherence, Aggregation, In vitro, Lactobacilli, PCR, Vaginal tract
  • Rashid Saif*, Ali Awan, Leslie Lyons, Barbara Gandolfi, Muhammad Tayyab, Masroor Ellahi Babar, Asim Mahmood, Zia Ullah, Muhammad Wasim Pages 142-152
    Background
    Molecular marker based cancer diagnosis gaining more attention in the current genomics era. So, Hspb1 and Tp53 gene characterization and their mRNA expression might be helpful in diagnosis andprognosis ofcat mammary adenocarcinoma. It will also add informationin comparative cancer genetics and genomics.
    Objectives
    Eight tumors of Siamese cats were analyzed to ascertain germ-line and tissue-specific somatic DNA variationsof Hspb1 and Tp53 genes alongwith the ectopic differential expressionin tumorous and normal tissueswere also analyzed.
    Materials And Methods
    Tumorous tissues and peripheral blood from mammary adenocarcinoma affected Siamese cats were collected from the Pet center-UVAS. DNA and RNA were extracted from these tissues to analyze the Hspb1 and Tp53 DNA variants and ectopic expression of their mRNA within cancerous and normal tissues.
    Results
    Exon 1 and 3 revealed as hotspots in Hspb1 gene. The 5´UTR region of the exon1 bearsix mutation including 3 transitions, 2 transversion and one heterozygous synonymous transversion in two samples at locus c.34C>C/A. Exon 3 has 1 transversion at c.773A>A/T, 3´UTR of this exon harbor two point mutationsat 1868A>T and 2193C>T loci. Intron 2 has two alterations at 1490C>C/T and GTCT4del at 1514. Overall up-regulationof Hspb1 gene was observed. While exons 3, 4 and 7 of Tp53 harbor asingle variationat c.105A>A/G, c.465T>T/C and c.859G>T respectively. The locus c.1050G>G/A in exon 9 is a heterozygous (G/A) in 3 samples and homozygous (G) in 2 other tumours. Introns 3, 5, 7 and 9 harbor 3, 4, 2 and 7 altered loci respectively. Sixty percentof cancers showed up-regulated trend of Tp53 gene.
    Conclusions
    Tumor specific mutations and ectopic expression of Hspb1 and Tp53 genes might be helpful in the diagnosis of the mammary lesions and endorse their involvement in cat mammary neoplasm.
    Keywords: Cat mammary tumor, Hsp27 mutation, expression, Tp53 mutation, expression