The efficient generation of cell lines from bovine parthenotes

The generation of embryonic stem cell (ESC) lines from parthenogenetically activated oocytes can provide transplantable cells, which are immunocompatible for the oocyte donors as well as an invaluable tool for genetic engineering and epigenetic studies. We report the efficient isolation of eight putative bovine parthenogenetic embryonic stem cell (bpESC) lines from 15 in vitro produced parthenotes. Five of these cell lines were maintained for more than 15 passages (>140 days) and analyzed. The cells displayed typical ESC morphology, stained positive for alkaline phosphate by histochemical staining, expressed Oct4, Nanog, and either stage-specific embryonic antigens, SSEA1, or SSEA4, detected by immunofluorescence staining. RT-PCR analysis of the cells demonstrated expression of Oct4, Rex1, SSEA1, and ALP. All the cell lines except one had a normal karyotype of 60, XX. The cells differentiated in suspension culture to form embryoid bodies (EBs) expressing markers of the three embryonic germ layers as assessed by RT-PCR. In conclusion, we report efficient derivation of putative ESCs from bovine parthenogenetic embryos. The cells express pluripotent markers, have the ability to form EBs, and differentiate into cells of the three embryonic germ layers. This is the first report of characterized putative parthenogenetic bovine ESC lines

Induction of Pluripotency in Adult Equine Fibroblasts without c-MYC

Despite tremendous efforts on isolation of pluripotent equine embryonic stem (ES) cells, to date there are few reports about successful isolation of ESCs and no report of in vivo differentiation of this important companion species. We report the induction of pluripotency in adult equine fibroblasts via retroviral transduction with three transcription factors using OCT4, SOX2, and KLF4 in the absence of c-MYC. The cell lines were maintained beyond 27 passages (more than 11 months) and characterized. The equine iPS (EiPS) cells stained positive for alkaline phosphatase by histochemical staining and expressed OCT4, NANOG, SSEA1, and SSEA4. Gene expression analysis of the cells showed the expression of OCT4, SOX2 NANOG, and STAT3. The cell lines retained a euploid chromosome count of 64 after long-term culture cryopreservation. The EiPS demonstrated differentiation capacity for the three embryonic germ layers both in vitro by embryoid bodies (EBs) formation and in vivo by teratoma formation. In conclusion, we report the derivation of iPS cells from equine adult fibroblasts and long-term maintenance using either of the three reprogramming factors

Unravelling evolution of Nanog, the key transcription factor involved in self-renewal of undifferentiated embryonic stem cells, by pattern recognition in nucleotide and tandem repeats characteristics

Nanog, an important transcription factor in embryonic stem cells (ESC), is the key factor in maintaining pluripotency to establish ESC identity and has the ability to induce embryonic germ layers. Nanog is responsible for self-renewal and pluripotency of stem cells as well as cancer invasiveness, tumor cell proliferation, motility and drug-resistance. Understanding the underlying mechanisms of Nanog evolution and regulation can lead to future advances in treatment of cancers. Recent integration of machine learning models with genetics has provided a powerful tool for knowledge discovery and uncovering evolutionary pathways. Herein, sequences of 47 Nanog genes from various species were extracted and two datasets of features were computationally extracted from these sequences. At the first dataset, 76 nucleotide acid attributes were calculated for each Nanog sequence. The second dataset was prepared based on the 10,480 repeated nucleotide sequences (from 5 to 50. bp lengths). Then, various data mining algorithms such as decision tree models were applied on these datasets to find the evolutionary pathways of Nanog diversion. Attribute weighting models were highlighted features such as the frequencies of AA and GC as the most important genomic features in Nanog gene classification and differentiation. Similar findings were obtained by tree induction algorithms. Results from the second database showed that some short sequence strings, such as ACTACT, TCCTGA, CCTGA, GAAGAC, and TATCCC can be effectively used to identify Nanog genes in various species. The outcomes of this study, for the first time, unravels the importance of particular genomic features in Nanog gene evolution paving roads toward better understanding of stem cell development and human targeted disorder therapy. © 2015 Elsevier B.V

Bidirectional and Opposite Effects of Naive Mesenchymal Stem Cells on Tumor Growth and Progression

Cancer has long been considered as a heterogeneous population of uncontrolled proliferation of different transformed cell types. The recent findings concerning tumorigeneses have highlighted the fact that tumors can progress through tight relationships among tumor cells, cellular, and non-cellular components which are present within tumor tissues. In recent years, studies have shown that mesenchymal stem cells (MSCs) are essential components of non-tumor cells within the tumor tissues that can strongly affect tumor development. Several forms of MSCs have been identified within tumor stroma. Naïve (innate) mesenchymal stem cells (N-MSCs) derived from different sources are mostly recruited into the tumor stroma. N-MSCs exert dual and divergent effects on tumor growth through different conditions and factors such as toll-like receptor priming (TLR-priming), which is the primary underlying causes of opposite effects. Moreover, MSCs also have the contrary effects by various molecular mechanisms relying on direct cellto- cell connections and indirect communications through the autocrine, paracrine routes, and tumor microenvironment (TME). Overall, cell-based therapies will hold great promise to provide novel anticancer treatments. However, the application of intact MSCs in cancer treatment can theoretically cause adverse clinical outcomes. It is essential that to extensively analysis the effective factors and conditions in which underlying mechanisms are adopted by MSCs when encounter with cancer. The aim is to review the cellular and molecular mechanisms underlying the dual effects of MSCs followed by the importance of polarization of MSCs through priming of TLRs

Enhancement of anticancer activity by silibinin and paclitaxel combination on the ovarian cancer

Background: Ovarian carcinoma is the most lethal cancer among all gynaecological malignancies. One of the most chemotherapy drugs used for ovarian cancer is paclitaxel which induces apoptosis. Paclitaxel has been used for many years. Similar to the most cancers this responds to chemotherapy initially but in a long run, drug resistance happens which fails the treatment procedure. Combination of chemotherapy drugs has been suggested to deal with this issue. Silibinin, a plant extraction, has been used from ancient time in traditional medicine and identified to have powerful antioxidant activity. Aim: The aim of this study was to examine the effect of paclitaxel and silibinin combination on SKOV-3 cancer cell line. Materials and methods: The human epithelial ovarian cancer cell line, SKOV-3, was cultured and treated with paclitaxel, silibinin and paclitaxel plus silibinin for 48 hours. MTT assay was carried out to determine cell viability. For apoptotic process, we used real-time PCR to study P53 and P21 genes expression after drug treatment and network analysis was performed using Pathway Studio web tool (Elsevier). Results: Cell growth was inhibited considerably (p <.05) by combination of paclitaxel and silibinin after 48 hours of treatment. Also silibinin and paclitaxel combination induced apoptosis in SKOV-3 cells. Expression analysis by real-time PCR showed the significant up-regulation of two tumour suppressor genes, P53 and P21 in response to combination of silibinin and paclitaxel. In addition, computational network analysis demonstrated the crosstalk between paclitaxel, silibinin and ovarian cancer. Conclusions: Our results showed that combination of chemotherapy drugs of silibinin and paclitaxel can be more efficient in treatment of ovarian cancer cells. © 2017, © 2017 Informa UK Limited, trading as Taylor & Francis Group

Enhancement of anticancer activity by silibinin and paclitaxel combination on the ovarian cancer

Background: Ovarian carcinoma is the most lethal cancer among all gynaecological malignancies. One of the most chemotherapy drugs used for ovarian cancer is paclitaxel which induces apoptosis. Paclitaxel has been used for many years. Similar to the most cancers this responds to chemotherapy initially but in a long run, drug resistance happens which fails the treatment procedure. Combination of chemotherapy drugs has been suggested to deal with this issue. Silibinin, a plant extraction, has been used from ancient time in traditional medicine and identified to have powerful antioxidant activity. Aim: The aim of this study was to examine the effect of paclitaxel and silibinin combination on SKOV-3 cancer cell line. Materials and methods: The human epithelial ovarian cancer cell line, SKOV-3, was cultured and treated with paclitaxel, silibinin and paclitaxel plus silibinin for 48 hours. MTT assay was carried out to determine cell viability. For apoptotic process, we used real-time PCR to study P53 and P21 genes expression after drug treatment and network analysis was performed using Pathway Studio web tool (Elsevier). Results: Cell growth was inhibited considerably (p <.05) by combination of paclitaxel and silibinin after 48 hours of treatment. Also silibinin and paclitaxel combination induced apoptosis in SKOV-3 cells. Expression analysis by real-time PCR showed the significant up-regulation of two tumour suppressor genes, P53 and P21 in response to combination of silibinin and paclitaxel. In addition, computational network analysis demonstrated the crosstalk between paclitaxel, silibinin and ovarian cancer. Conclusions: Our results showed that combination of chemotherapy drugs of silibinin and paclitaxel can be more efficient in treatment of ovarian cancer cells. © 2017, © 2017 Informa UK Limited, trading as Taylor & Francis Group

Legionella Pneumophila and Dendrimers-Mediated Antisense Therapy

Finding novel and effective antibiotics for treatment of Legionella disease is a challenging field. Treatment with antibiotics usually cures Legionella infection; however, if the resultant disease is not timely recognized and treated properly, it leads to poor prognosis and high case fatality rate. Legionella pneumophila DrrA protein (Defects in Rab1 recruitment protein A)/also known as SidM affects host cell vesicular trafficking through modification of the activity of cellular small guanosine triphosphatase )GTPase( Rab (Ras-related in brain) function which facilitates intracellular bacterial replication within a supporter vacuole. Also, Legionella pneumophila LepA and LepB (Legionella effector protein A and B) proteins suppress host-cell Rab1 protein's function resulting in the cell lysis and release of bacteria that subsequently infect neighbour cells. Legionella readily develops resistant to antibiotics and, therefore, new drugs with different modes of action and therapeutic strategic approaches are urgently required among antimicrobial drug therapies;gene therapy is a novel approach for Legionnaires disease treatment. On the contrary to the conventional treatment approaches that target bacterial proteins, new treatment interventions target DNA (Deoxyribonucleic acid), RNA (Ribonucleic acid) species, and different protein families or macromolecular complexes of these components. The above approaches can overcome the problems in therapy of Legionella infections caused by antibiotics resistance pathogens. Targeting Legionella genes involved in manipulating cellular vesicular trafficking using a dendrimer-mediated antisense therapy is a promising approach to inhibit bacterial replication within the target cells

Cryopreservation and long-term maintenance of bovine embryo-derived cell lines

The aim of this study was to develop methods for cryopreservation and long-term maintenance of putative bovine embryonic stem cells (ESCs). Putative bovine ESC (bESC) lines (n≤3) isolated in conventional medium were used to compare slow-freezing and vitrification. After warming, vitrified cells (96.9%) demonstrated significantly (