On Monday, June 1, SIVB’s In Vitro Animal Cell Sciences Section (IVACS) held a Student and Post-Doctoral Oral Presentation Competition that was moderated by Dr. Kolla Kristjansdottir (Midwestern University) and Dr. Addy Alt-Holland (Tufts University). Many abstracts have been submitted to be considered for this competition and the top contestants were chosen to present their research based on the quality and scientific merit of their abstracts and accomplishments. The three presentations were highly professional and the level of the presented research work was impressive and stimulating. Follwing their presentations, the participants received many questions from both the audience and the judges. The panel of IVACS experts that evaluated the contestants included Dr. Mae Ciancio and Dr. Michael J. Fay (Midwestern University), Dr. Barbara Doonan (New York Medical College), Dr. John Harbell (JHarbell Consulting LLC), Dr. Brad L. Upham (Michigan State University), Dr. Michael Dame (University of Michigan) and the moderators Dr. Kolla Kristjansdottir and Dr. Addy Alt-Holland. During the IVACS Business Meeting we were happy to announce that Allison Songstad who is a graduate research assistant at the University of Iowa won the 1st place award, Alexandra J. Cho who is a graduate student at Cornell University won the 2nd place award, and Razi Ahmad who is a research scholar at Jamia Millia Islamia won the 3rd place award. Certificates and monetary awards were presented to these students at that meeting. This Oral Presentation Competition provides an invaluable opportunity for students and Post-Doctoral candidates to present their research work to an expert audience and enrich their experience and achievements. We would like to encourage all qualified individuals to submit their research abstracts to the 2016 competition.
Submitted by Addy Alt-Holland and Kolla Kristjansdottir
Generation of Induced Pluripotent Stem Cell-derived Choroidal Endothelial Cells for the Treatment of AMD
Age-related macular degeneration (AMD), the most common cause of incurable blindness in the western world, involves the dysfunction and eventual death of choroidal endothelial (CEC), retinal pigment epithelial (RPE), and photoreceptor cells. Induced pluripotent stem cell (iPSC)-based strategies designed to replace both photoreceptor and RPE cells are currently a major scientific focus. For some patients, success of these approaches may also require replacement of CECs. The purpose of this study was to generate CECs from two Tie2 GFP iPSC reporter lines to develop efficient differentiation and transplantation protocols. Dermal fibroblasts from the Tie2 GFP mouse (carrying a GFP reporter gene under the control of the EC-specific Tie2 promoter) were isolated and reprogrammed into two iPSC lines via viral transduction of the Yamanaka factors Oct4, Sox2, Klf4, and c-Myc. iPSC potency was characterized via RT-PCR, immunocytochemistry (ICC), and teratoma formation assays. iPSC-CECs were differentiated using a co-culture method with either the RF6A CEC line or primary mouse CECs. iPSC-CECs from both lines were characterized via RT-PCR and ICC for various EC- and CEC-specific markers. The two iPSC-derived CEC lines expressed the EC markers CD31, CD34, Tie2, VE-Cadherin, and VWF, as determined by RT-PCR, expressed the CEC-specific markers carbonic anhydrase IV and ICAM1, as determined by RT-PCR and ICC, and were morphologically indistinguishable from native CECs that expressed ZO-1 and VE-Cadherin in cell-cell adhesions. Thus, we successfully generated murineiPSCs that can be efficiently differentiated into CECs. This work has paved the way for future studies focused on investigating AMD pathophysiology and CEC replacement.
Allison Songstad, University of Iowa, University of Iowa, Stephen A. Wynn Institute for Vision Research, Carver College of Medicine, Department of Ophthalmology, 375 Newton Rd., 4156 MERF, Iowa City, 52242. In Vitro Cellular and Developmental Biology, 51:S26-27, 2015
Development of Tools to Understand the Inverse Association Between Total Plasma Adiponectin and Liver NTMT1
Adiponectin is a highly abundant circulating peptide hormone secreted by adipocytes with known anti-inflammatory and insulin-sensitizing roles (Iwabu et al. 2010). Its role in increasing fatty acid oxidation in the liver is mediated through the activation of AMPK, which in turn regulates DNMT1 deacetylation downstream (Yamauchi et al. 2002). DNMT1 is responsible for maintaining DNA methylation patterns and normal liver function (Raggi et al. 2014). Our group is currently working on the development of a human in vitro model to study epigenetic dysregulations. Consequently, there is a need to develop tools for the assessment of adiponectin in secretory fluids. This research is part of the IRB approved clinical trial GHS #1207-27. We found an inverse association between levels of total circulating adiponectin and levels of liver DNMT1 in obese patients (r = -0.7614, p = 0.0002, n = 18). Futhermore, the highest levels of adiponectin in this cohort of patients (mean BMI 44) were accompanied by the lowest liver DNMT1 levels. To the best of our knowledge, this is the first report of the association between adiponectin and liver DNMT1 in human subjects. Currently, we work towards the development of tools for the assessment of adiponectin in vivo and in vitro.
Alexandra H. Cho, Cornell University, Department of Biomedical Engineering, Cornell University and 2Guthrie Clinic, Sayre, PA. In Vitro Cellular and Developmental Biology, 51:S26, 2015
Evaluation of Green Synthesis TiO2 Nanoparticles as Antibacterial Agents
Nanotechnology is gaining momentum due to its diverse application in various fields of science. We have synthesized TiO2 nanoparticles by two approaches by using alpha amylase enzyme as the sole reducing and capping agent and using lactobacillus sp. The biosynthesized nanoparticles were characterized by X-ray diffraction (XRD) and transmission electron microscopic (TEM) methods. The XRD data confirms the monophasic crystalline nature of the nanoparticles formed. TEM data shows that the particle size synthesized by alpha amylase is varies in the range of 30-70 nm with an average size of 50 nm, whereas particle size synthesized by lactobacillus sp. is in the range of 50-100 nm with an average size of 75nm. FTIR peaks indicate that the biomolecules is responsible for the synthesis and stabilization of TiO2 nanoparticles. The nanoparticles were further investigated for their antibacterial effect on Staphylococcus aureus and Escherichia coli. The minimum inhibitory concentration value of the biologically synthesized TiO2 nanoparticles were found to be 62.50 µg/ml for both the bacterial strains. The inhibition was further confirmed using disc diffusion assay. It is evident from the zone of inhibition that TiO2 nanoparticles possess potent bactericidal activity. Further, growth curve study of S. aureus and E. coli in the presence of TiO2 nanoparticles shows effect of inhibitory concentration of TiO2 nanoparticles against S. aureus and E. coli. TEM investigation confirmed that nanoparticles are disrupting the bacterial cell wall. This study indicates that the biologically synthesized nanoparticles can be used as effective antibacterial agents.
Razi Ahmad, Jamia Millia Islamia, Department of Biosciences, Jamia Millia Islamia, New Delhi-110025, INDIA. In Vitro Cellular and Developmental Biology, 51:S26, 2015