The members of the laboratory group that the performed the work described in this manuscript are, from left to right (front row, seated) Hong-Bing Chen, Eva Zipkin, Nathan Moore, (back row, standing) Kyle Draheim, Dr. Stephen Lyle, Dr. Zhiru Guo, Kathryn Bomba and Michael Roche.
Multi-potentiality of a New Immortalized Epithelial Stem Cell Line Derived from Human Hair Follicles
Research in this laboratory focuses on the molecular and cellular characterization of adult epithelial stem cells and pathways which lead to cancer from stem cells. The stem cells of human and murine skin reside in a specific area (niche) of the hair follicle called the “bulge”. These cells have stem cell properties of self-renewal, slow-cycling and multi-potentiality within skin, giving rise to epidermis, sebaceous gland and hair follicle. These bulge cells are also responsible for regeneration of the hair follicle during hair cycling. A number of differentially expressed genes within the stem cell compartment of skin have recently been identified, however the functions of many of these genes in mediating stem cell properties such as self-renewal, growth, differentiation, adhesion and migration have not been explored. In addition, studies of altered signaling pathways that may lead to malignant transformation of stem cells have been hampered by a lack of well-characterized adult stem cell lines. This study showed that an immortalized cell line derived from the adult human hair follicle bulge (Tel-E6E7) has the same phenotype as native stem cells and can be passaged indefinitely. Unlike other skin cell lines such as HaCat cells, the Tel-E6E7 cells maintain their multi-potentiality in vitro by responding to inductive signals to undergo a hair follicle phenotype, as well as differentiate towards epidermis and sebocytes. The cells express some of the stem cell markers found in vivo and possess similar characteristics of clonogenicity, migration and adhesion. The Tel-E6E7 cell line should provide a reproducible in vitro model for studies of stem cell lineage determination and differentiation as well as other stem cell properties. In addition, molecular mechanisms that may lead to adult stem cell transformation can be readily investigated.
Cecilia Roh, Michael Roche, Zhiru Guo, Christos Photopoulos, Qingfeng Tao and Stephen Lyle. Multi-potentiality of a new immortalized epithelial stem cell line derived from human hair follicles, In Vitro Cellular & Developmental Biology-Animal 44:236-244, 2008.
Electrophysiological and Iimmunocytochemical Characterization of DRG Neurons on an Organosilane Surface in Serum-free Medium
One of the primary research projects in the Hybrid Systems Laboratory is focused on developing a bio-engineered in vitro model of the stretch reflex arc. Successful creation of a functional model of the reflex arc would enable a new system to develop effective therapies for injury and diseases in the spinal cord, such as ALS, muscle wasting diseases, such as muscular dystrophy, as well as a better understanding of neuropathic pain. The project utilizes a multi-disciplinary approach, drawing from the areas of cell biology, nano-surface chemistry and tissue engineering. The stretch reflex arc model integrates advances in serum-free, defined medium development, a non-biological growth substrate and bio-microelectromechanical systems (bio-MEMS) cantilever arrays. These approaches provide a highly controlled system for engineering and integrating the components of the stretch reflex arc and support the analysis of signal conversion from mechanical muscle stretch to electrical impulses and finally back to mechanical muscle contraction. Currently, we are using various aspects of this system for research into neuromuscular junction formation, intrafusal muscle development and for new approaches for regenerative medicine and prosthetic device design. In our recently published research, rat sensory neurons were grown on the biomimetic substrate DETA using defined serum-free conditions. Under these conditions, all three types of sensory neurons were shown to express specific trk receptor groups as well as normal neuronal cytoskeletal proteins. The neurons also demonstrated normal inward Na+ and outward K+ currents and fired both single and repetitive action potentials. This research represents the successful growth, morphology and functional activity of one component of sensory circuit of the stretch reflex arc.
Jie Liu,John W Rumsey,Mainak Das, Peter Molnar,Cassie Gregory, Lisa Riedel, and James J Hickman. Electrophysiological and immunocytochemical characterization of DRG neurons on an organosilane surface in serum-free medium, In Vitro Cellular & Developmental Biology-Animal 44:162-168, 2008.
Evaluation of methods for celery (Apium Graveolens L.) transformation using Agrobacterium tumefaciens and the bar gene as selectable marker
The Plant Biotechnology Resource and Outreach Center (PBROC) (former Plant Transformation Center) of Michigan State University is dedicated to developing efficient gene transfer and regeneration systems for specialty and orphan crops such as blueberry, cherry, dry beans, and celery. The center also provides contract services in molecular fingerprinting and presents training in environmental biosafety. The work described herein focused on developing a reliable transformation system for celery using Agrobacterium tumefaciens and the bialaphos resistance (bar) gene as selectable marker. A. tumefaciens strains EHA105 and GV3101, each with the bar gene under the promoters nopaline synthase (nos) (pGPTV-BAR) or cauliflower mosaic virus (CaMV) 35S (pDHB321.1), were used for transformation of two celery cultivars XP85 and XP166. Callus selection (CS) and the flamingo-bill explant (FB) methods were evaluated for efficacy in production of transgenic plants. Using the CS protocol, it took 5-6 months total time to obtain transgenic plants. Conversely, the FB protocol yielded putative transgenic celery plants in just 6 weeks. Stable integration of the bar gene with 1-2 copies were confirmed by Southern blot analysis in primary transformants derived by both methods. Herbicide assays on primary transformants indicated a range of low to high tolerance to 100 or 300 mg·l-1 glufosinate. Progeny analysis by polymerase chain reaction (PCR) showed stable Mendelian inheritance of the bar gene.
Andrey V Loskutov, Guo-Qing Song, and Kenneth Sink. Evaluation of methods for celery (Apium Graveolens L.) transformation using Agrobacterium tumefaciens and the bar gene as selectable marker, In Vitro Cellular & Developmental Biology-Plant 44:239-245, 2008.
