Incorporation of SPION-casein core-shells into silk-fibroin nanofibers for cardiac tissue engineering

Mimicking the structure of extracellular matrix (ECM) of myocardium is necessary for fabrication of functional cardiac tissue. The superparamagnetic iron oxide nanoparticles (SPIONs, Fe₃O₄), as new generation of magnetic nanoparticles (NPs), are highly intended in biomedical studies. Here, SPION NPs (1 wt%) were synthesized and incorporated into silk-fibroin (SF) electrospun nanofibers to enhance mechanical properties and topography of the scaffolds. Then, the mouse embryonic cardiac cells (ECCs) were seeded on the scaffolds for in vitro studies. The SPION NPs were studied by scanning electron microscope (SEM), X-ray diffraction (XRD), and transmission electron microscope (TEM). SF nanofibers were characterized after incorporation of SPIONs by SEM, TEM, water contact angle measurement, and tensile test. Furthermore, cytocompatibility of scaffolds was confirmed by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. SEM images showed that ECCs attached to the scaffolds with elongated morphologies. Also, the real-time PCR and immunostaining studies approved upregulation of cardiac functional genes in ECCs seeded on the SF/SPION-casein scaffolds including GATA-4, cardiac troponin T, Nkx 2.5, and alpha-myosin heavy chain, compared with the ones in SF. In conclusion, incorporation of core-shells in SF supports cardiac differentiation, while has no negative impact on ECCs' proliferation and self-renewal capacity.     

Nanoparticles and cancer therapy: Perspectives for application of nanoparticles in the treatment of cancers

Rapid growth in nanotechnology toward the development of nanomedicine agents holds massive promise to improve therapeutic approaches against cancer. Nanomedicine products represent an opportunity to achieve sophisticated targeting strategies and multifunctionality. Nowadays, nanoparticles (NPs) have multiple applications in different branches of science. In recent years, NPs have repetitively been reported to play a significant role in modern medicine. They have been analyzed for different clinical applications, such as drug carriers, gene delivery to tumors, and contrast agents in imaging. A wide range of nanomaterials based on organic, inorganic, lipid, or glycan compounds, as well as on synthetic polymers has been utilized for the development and improvement of new cancer therapeutics. In this study, we discuss the role of NPs in treating cancer among different drug delivery methods for cancer therapy.     

Slow Regulated Release of H2S Inhibits Oxidative Stress Induced Cell Death by Influencing Certain Key Signaling Molecules

Hydrogen sulphide (H₂S) is one of three gaseous signaling molecules after nitric oxide and carbon monoxide. Various H₂S donor compounds have been synthesized to study its physiological function. Among these compounds sodium hydrosulphide (NaHS), a donor of releasing H₂S rapidly have shown to be protective in certain neuronal cell line but several in vivo studies have generated conflicting data. Furthermore several slow releasing H₂S donors have been shown to have positive effects on cells in culture. The intracellular concentration of H₂S and hence its rate of production may be a factor in keeping the balance between its neuroprotective and toxic effects. The present study was undertaken to deduce how a rapid releasing H₂S donor (NaHS) as opposed to a slow releasing donor (ADTOH), affect oxidative stress related intracellular components and survival of RGC-5 cells. It was concluded that when RGC-5 cells are exposed to the toxic effects of glutamate in combination with buthionine sulfoxime (Glu/BSO), ADTOH was more efficacious in inhibiting apoptosis, scavenging reactive oxygen species (ROS), stimulation of glutathione (GSH) and gluthathione-S-transferase (GST). Western blot and qPCR analysis showed ADTOH increased the levels of Nrf2, HO-1, PKCα, p-Akt, Bcl-2 and XIAP but caused a decrease of Nfκβ and xCT greater than NaHS. This study is first to compare the efficacy of two H₂S donor drugs as potential neuroprotectants and demonstrate that slow regulated release of H₂S to cell culture can be more beneficial in inhibiting oxidative stress induced cell death.     

Efficient growth inhibition of EGFR over-expressing tumor cells by an anti-EGFR nanobody

Epidermal growth factor receptor (EGFR) is deemed to be one of the main molecular targets for diagnosis and treatment of cancer. It has been identified that EGFR involves in pathogenesis of some forms of human cancers. Monoclonal antibodies targeting EGFR could control the tumor cell growth, proliferation, and apoptosis by suppressing the signal transduction pathways. Nanobodies can be regarded as the smallest intact antigen binding fragments, derived from heavy chain-only antibodies existing in camelids. Here, we describe the identification of an EGFR-specific nanobody, referred to as OA-cb6, obtained from immunized camel with a cell line expressing high levels of EGFR. Utilizing flow cytometry (FACS) and blotting methods, we demonstrated that OA-cb6 nanobody binds specifically to EGFR expressing on the surface of A431 cells. In addition, OA-cb6 nanobody potently causes the inhibition of EGFR over expression, cell growth and proliferation. The antibody fragments can probably be regarded as worthwhile binding block for further rational design of anti-cancer therapy.     

Probing the interaction of silver nanoparticles with tau protein and neuroblastoma cell line as nervous system models

Interestingly pharmaceutical sciences are using nanoparticles (NPs) to design and develop nanomaterials-based drugs. However, up to recently, it has not been well realized that NPs themselves may impose risks to the biological systems. In this study, the interaction of silver nanoparticles (AgNPs) with tau protein and SH-SY5Y neuroblastoma cell line, as potential nervous system models, was examined with a range of techniques including intrinsic fluorescence spectroscopy, circular dichroism (CD) spectroscopy, 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and acridine orange/ethidium bromide (AO/EB) dual staining method. Fluorescence study showed that AgNPs with a diameter of around 10–20 nm spontaneously form a static complex with tau protein via hydrogen bonds and van der Waals interactions. CD experiment revealed that AgNPs did not change the random coil structure of tau protein. Moreover, AgNPs showed to induce SH-SY5Y neuroblastoma cell mortality through fragmentation of DNA which is a key feature of apoptosis. In conclusion, AgNPs may induce slight changes on the tau protein structure. Also, the concentration of AgNPs is the main factor which influences their cytotoxicity. Since, all adverse effects of NPs are not well detected, so probably additional more specific testing would be needed.     

Comparative immunomodulatory properties of adipose‐derived mesenchymal stem cells conditioned media from BALB/c,C57BL/6, and DBA mouse strains

Adipose tissue‐derived mesenchymal stem cells (AD‐MSCs) have been shown to be capable of differentiating into multiple cell type and exert immunomodulatory effects. Since the selection of ideal stem cell is apparently crucial for the outcome of experimental stem cell therapies, therefore, in this study we compared AD‐MSCs conditioned media (CM) from BALB/c, C57BL/6, and DBA mouse strains. No significant difference was found in the morphology, cell surface markers, in vitro differentiation and proliferation potentials of AD‐MSCs isolated from C57BL/6, BALB/c, and DBA mice. The immunological assays showed some variation among the strains in the cytokines, nitric oxide (NO), and indoleamine 2,3‐dioxygenase (IDO) production and immunomodulatory effects on splenocytes functions. Our results indicated a suppression of splenocytes proliferation in the presence of AD‐MSC CM from the three inbred mouse strains. However, BALB/c CM exerted a higher suppression of splenocytes proliferation. AD‐MSCs isolated from C57BL/6 and BALB/c mice produced higher levels of TGF‐β than those from DBA mice. Furthermore, IL‐17 and IDO production was higher in AD‐MSCs isolated from BALB/c mice. Our results indicated an increased production of TGF‐β, IL‐4, IL‐10, NO, and IDO by splenocytes in response to CM from BALB/c AD‐MSCs. In conclusion, our results showed that the immunomodulatory properties of mouse AD‐MSCs is strain‐dependent and this variation should be considered during selection of appropriate stem cell source for in vivo experiments and stem cell therapy strategies. J. Cell. Biochem. 114: 955–965, 2013. © 2012 Wiley Periodicals, Inc.     

Normalization of miRNA qPCR high-throughput data: a comparison of methods

Low-density quantitative real-time PCR (qPCR) arrays are often used to profile expression patterns of microRNAs in various biological milieus. To achieve accurate analysis of expression of miRNAs, non-biological sources of variation in data should be removed through precise normalization of data. We have systematically compared the performance of 19 normalization methods on different subsets of a real miRNA qPCR dataset that covers 40 human tissues. After robustly modeling the mean squared error (MSE) in normalized data, we demonstrate lower variability between replicates is achieved using various methods not applied to high-throughput miRNA qPCR data yet. Normalization methods that use splines or wavelets smoothing to estimate and remove Cq dependent non-linearity between pairs of samples best reduced the MSE of differences in Cq values of replicate samples. These methods also retained between-group variability in different subsets of the dataset.     

Motor Domain Phosphorylation Modulates Kinesin-1 Transport

Disruptions in microtubule motor transport are associated with a variety of neurodegenerative diseases. Post-translational modification of the cargo-binding domain of the light and heavy chains of kinesin has been shown to regulate transport, but less is known about how modifications of the motor domain affect transport. Here we report on the effects of phosphorylation of a mammalian kinesin motor domain by the kinase JNK3 at a conserved serine residue (Ser-175 in the B isoform and Ser-176 in the A and C isoforms). Phosphorylation of this residue has been implicated in Huntington disease, but the mechanism by which Ser-175 phosphorylation affects transport is unclear. The ATPase, microtubule-binding affinity, and processivity are unchanged between a phosphomimetic S175D and a nonphosphorylatable S175A construct. However, we find that application of force differentiates between the two. Placement of negative charge at Ser-175, through phosphorylation or mutation, leads to a lower stall force and decreased velocity under a load of 1 piconewton or greater. Sedimentation velocity experiments also show that addition of a negative charge at Ser-175 favors the autoinhibited conformation of kinesin. These observations imply that when cargo is transported by both dynein and phosphorylated kinesin, a common occurrence in the cell, there may be a bias that favors motion toward the minus-end of microtubules. Such bias could be used to tune transport in healthy cells when properly regulated but contribute to a disease state when misregulated.     

Validation and fine mapping of lyc12.1, a QTL for increased tomato fruit lycopene content

Lycopene content is a key component of tomato (Solanum lycopersicum L.) fruit quality, and is a focus of many tomato-breeding programs. Two QTLs for increased fruit lycopene content, inherited from a high-lycopene S. pimpinellifolium accession, were previously detected on tomato chromosomes 7 and 12 using a S. lycopersicum × S. pimpinellifolium RIL population, and were identified as potential targets for marker-assisted selection and positional cloning. To validate the phenotypic effect of these two QTLs, a BC₂ population was developed from a cross between a select RIL and the S. lycopersicum recurrent parent. The BC₂ population was field-grown and evaluated for fruit lycopene content using HPLC. Statistical analyses revealed that while lyc7.1 did not significantly increase lycopene content in the heterozygous condition, individuals harboring lyc12.1 in the heterozygous condition contained 70.3 % higher lycopene than the recurrent parent. To eliminate the potential pleiotropic effect of fruit size and minimize the physical size of the lyc12.1 introgression, a marker-assisted backcross program was undertaken and produced a BC₃S₁ NIL population (n = 1,500) segregating for lyc12.1. Lycopene contents from lyc12.1 homozygous and heterozygous recombinants in this population were measured and lyc12.1 was localized to a 1.5 cM region. Furthermore, we determined that lyc12.1 was delimited to a ~1.5 Mb sequence of tomato chromosome 12, and provided some insight into potential candidate genes in the region. The derived sub-NILs will be useful for transferring of lyc12.1 to other tomato genetic backgrounds and for further fine-mapping and cloning of the QTL.     

Investigation of cytocrom c oxidase gene subunits expression on the Multiple sclerosis

INTRODUCTION:

Multiple sclerosis (MS) is an autoimmune inflamatory disease, which affects the (Central Nervous System) and leads to the destruction of myelin and atrophy of the axons. Genetic factors, in addition to environmental ones, seem to play a role in MS. Numerous studies have reported mitochondrial defects including a reduction in cytochrome c oxidase (COX) complex function related to the reduction of mitochondrial genes expression in the cortex tissue of patients with MS have been reported.

MATERIALS AND METHODS:

This study aimed to assess COX5B and COX2 genes expression in MS patients and compare it with normal subjects. We determine expression levels of genes COX5B and COX2, and also gene reference ß-actine using real–time polymerase chain reaction (RT-PCR) method. Data were obtained and obtained and standardized with the gene reference and were analyzed using independent sample t-test with SPSS and Excel programs.

RESULT AND DISCUSSION:

The resultshowed COX5B gene expression reduced significant in MS patients compared to normal subjects (P < −0.05) whereas, there was no significant difference in the COX2 gene expression between normal subjects and patients. Thus, it can be claimed that down-regulation of mitochondrial electron transport chain genes supported the hypothesis that hypoxia-like tissue injury in MS may be due to mitochondrial genes, different expression impairment.