Detection of Intracellular Adhesion (ica) Gene and Biofilm Formation Staphylococcus aureus Isolates from Clinical Blood Cultures

Background: In fact the biofilms are composed of bacterial cells living inmulticellular structures such as tissues and organs embedded within a self-produced matrix of extracellular polymeric substance (EPS). Ability to attach and biofilm formation are the most important virulence factors Staphylococcus aureus isolates. The aims of this study were to detect intracellular adhesion (ica) locus and its relation to the biofilm formation phenotype in clinical isolates of S. aureus isolated from bloodcultures. Methods: A total of 31 clinical S. aureus isolates were collected from Loghman Hospital of Tehran, Iran. In vitro biofilm formation ability was determined by microliter tissue culture plates. All clinical isolates were examined for determination the ica locus by using PCR method. Results: Twelve (38.7%) of the isolates were strong biofilm producers. The results showed that 18(80.6%) of the isolates carried icaD gene, whereas the prevalence of icaA, icaB and icaC were 51.6%, 45.1% and 77.4% respectively. Conclusions: S. aureus clinical isolates have different ability to form biofilm. This may be caused by the differences in the expression of biofilm related genes, genetic make-up and physiological conditions

Inhibition of breast cancer metastasis by using miR-31-mimic in cancer stem cell rich MDA-MB231 cell line

Background: Metastasis associated miRNA (metastamiR) opened a new field of anti-metastatic therapy which have a great potential of treatment for the most lethal aspect of cancer, metastasis. The pleiotropic nature of gene regulation exhibited by certain miRNAs that showed that miRNAs might be endowed with a capacity to function as crucial modulators of tumor metastasis. MiR-31 is a pleiotropic anti-metastatic miRNA whose expression decreased significantly in metastatic breast cancer cells. MiR-31 has multiple roles in metastasis cascade. Therefore, using the miR-31-restoration based therapy could be an efficient anti-metastatic strategy for cancer therapy. Methods: This research was performed from May 2014 to October 2015 in Tarbiat Modares University in Tehran, Iran. The double-strand oligo of mature miR-31 was cloned into pcDNA 6.2gw/EmGFP according to the manufacturer instruction. The MDA-MB231, MCF-7 breast cancer cell lines were cultured and their miRNAs have been extracted. The expression of miR-31 has been quantified by Real time-PCR be-fore transfection of construct contained miR-31 into two cell lines and in normal breast cells. Then the constructs contain miR-31 have been transfected in to two cell lines. The expression of miR-31 has been quantified after 48 hours. Scratch and invasion as-say have been carried out for assessing the level of migration and invasion. Results: The result of Real time-PCR before transfection of constructs contained miR-31 have been shown 4 fold and more than 100 fold reduction in expression of miR-31 in MCF-7 and MDA-MB231 respectively in comparison to miR-31 expression in nor-mal breast cells, but after transfection of miR-31 construct to MDA-MB231 the quan-tification of expression showed the significant increase in mir-31 expression and 20 fold reduction in invasive and 10 fold reduction in migratory characteristics of MDA-MB231 in comparison to MCF-7. Conclusion: Metastasis associated miRNA have been represented a promising candi-dates in the field of anti-metastatic therapy and miR-31 as a powerful member of this family can function very effectively in order to inhibit the metastasis and introduce the new possibility of metastasis inhibition

Comparison of the Ex Vivo Expansion of UCB-Derived CD34+ in 3D DBM/MBA Scaffolds with USSC as a Feeder Layer

  　 Objective(s): Ex vivo expansion of hematopoitic stem cells is an alternative way to increase umbilical cord blood (UCB)-CD34+ cells for bone marrow transplantation. For this purpose demineralized bone matrix (DBM) and mineralized bone allograft (MBA) as two scaffolds based on bone matrix and stem cell niche, were simultaneously used to enhance the effect of human mesenchymal progenitor cells (MPCs) - unrestricted somatic stem cells (USSCs) - as a feeder layer.   Materials and Methods: USSCs were isolated and characterized by morphological and immunological analysis then seeded on both scaffolds as a feeder layer. UCB-CD34+ were isolated by MACS method and were co-culture expanded by USSC in 3D and 2D environments. After 3 weeks expansion, cells were counted and were assessed by karyotype, flow cytometry, clonogenic activity, and long-term culture-initiating cells (LTC-IC). Results: Co-culture expansion in DBM and MBA was 29.22-fold and 27.77-fold, no significant differences in colony and LTC-IC were obtained. Maximum number of colonies belonged to the day 14 with the 73% CFU-GM (Colony Forming Unit- Granulocyte/Macrophage) in contrast to the day 0 which was BFU-E/CFU-E (Burst/Colony Forming Unit-Erythroid). Flow cytometry indicated that the percentage of CD34+ marker was decreased in USSC co-culture and the highest percentage was observed in simple 2D culture. Conclusion: Because of acid extraction in the DBM production process, mineral materials were removed and the protein background that was more flexible was presented. Therefore these results suggest that USSC-DBM can be a suitable ex vivo mimicry niche by intensifying of surface/volume ratio and supporting the stem cell differentiation and expansion