K. Sue O’Shea

Stem cell transplantation for auditory nerve replacement

The successful function of cochlear prostheses depends on activation of auditory nerve. The survival of auditory nerve neurons, however, can vary widely in candidates for cochlear implants and influence implant efficacy. Stem cells offer the potential for improving the function of cochlear prostheses and increasing the candidate pool by replacing lost auditory nerve. The first phase of studies for stem cell replacement of auditory nerve has examined the in vitro survival and differentiation as well as in vivo differentiation and survival of exogenous embryonic and tissue stem cells placed into scala tympani and/or modiolus. These studies are reviewed and new results on in vivo placement of B-5 mouse embryonic stem cells into scala tympani of the guinea pig cochleae with differentiation into a glutamatergic neuronal phenotype are presented. Research on the integration and connections of stem cell derived neurons in the cochlea is described. Finally, an alternative approach is considered, based on the use of endogenous progenitors rather than exogenous stem cells, with a review of promising findings that have identified stem cell-like progenitors in cochlear and vestibular tissues to provide the potential for auditory nerve replacement.

Lumen Formation Is an Intrinsic Property of Isolated Human Pluripotent Stem Cells

Summary We demonstrate that dissociated human pluripotent stem cells (PSCs) are intrinsically programmed to form lumens. PSCs form two-cell cysts with a shared apical domain within 20 hr of plating; these cysts collapse to form monolayers after 5 days. Expression of pluripotency markers is maintained throughout this time. In two-cell cysts, an apical domain, marked by EZRIN and atypical PKCζ, is surrounded by apically targeted organelles (early endosomes and Golgi). Molecularly, actin polymerization, regulated by ARP2/3 and mammalian diaphanous-related formin 1 (MDIA), promotes lumen formation, whereas actin contraction, mediated by MYOSIN-II, inhibits this process. Finally, we show that lumenal shape can be manipulated in bioengineered micro-wells. Since lumen formation is an indispensable step in early mammalian development, this system can provide a powerful model for investigation of this process in a controlled environment. Overall, our data establish that lumenogenesis is a fundamental cell biological property of human PSCs.

Thrombospondin expression in nerve regeneration I. Comparison of sciatic nerve crush, transection, and long-term denervation.

Patterns of expression of the extracellular matrix molecule thrombospondin (TSP) were examined during peripheral nerve regeneration following sciatic nerve crush or transection. In noninjured nerve, was present in the axoplasm, Schwann cells, endoneurium, and perineurium of the adult mouse sciatic nerve. Following nerve crush or nerve transection, levels of TSP rapidly increased distal to the trauma site. Elevated levels of TSP were present distal to regenerating axons, while expression gradually returned to normal proximal to the regenerating axons. When reinnervation was blocked, TSP levels remained high in the endoneurium in excess of 30 days, but TSP was absent by 60 days. Following reanastomosis of the proximal and distal segments after 60 days of denervation, TSP was re-expressed in the distal nerve stump. These results indicate that TSP, which is involved in neuronal migrations in the embryo and neurite outgrowth in vitro, appears to play a role in axonal regeneration in the adult peripheral nervous system.

Thrombospondin expression in nerve regeneration II. Comparison of optic nerve crush in the mouse and goldfish

Expression of the extracellular matrix molecule thrombospondin (TSP) was examined following retrobulbar crush injury of the goldfish and mouse optic nerve. TSP was present within the glia limitans and surrounding axon fascicles of the control normal goldfish optic nerve, but was absent from the normal mouse optic nerve. Following crush injury of the goldfish optic nerve, TSP expression increased dramatically along the path of regenerating axons and returned to near normal levels following axonal outgrowth. In contrast, during the unsuccessful attempt at regeneration following crush injury of the mouse optic nerve, TSP expression was present only in glial fibrillary acidic protein (GFAP)-negative, macrophage-rich regions distal to ganglion cell axons. These results indicate that TSP expression is increased in a temporal pattern along the path of regenerating goldfish optic nerve axons and therefore may be involved in successful central nervous system regeneration. The absence of TSP in the environment encountered by damaged mouse optic nerve axons may correlate with the lack of regeneration observed in the mouse optic nerve.

Gene expression alterations in bipolar disorder postmortem brains

Objectives:  Bipolar disorder (BD) is a mental illness of unknown neuropathology and has several genetic associations. Antipsychotics are effective for the treatment of acute mania, psychosis, or mixed states in individuals with BD. We aimed to identify gene transcripts differentially expressed in postmortem brains from antipsychotics‐exposed individuals with BD (hereafter the ‘exposed’ group), non‐exposed individuals with BD (hereafter the ‘non‐exposed’ group), and controls.

Neural Differentiation of Embryonic Stem Cells

Neural stem cells exist in the mammalian developing and adult nervous system during the neural differentiation of the embryonic stem cells. Stem cells are important cells for replacement therapy diseases. The interest in the potential of for the treatment of neurodegenerative diseases and brain injuries has substantially promoted research on neural stem cell self-renewal and differentiation. General chapters of this review will deal with the history and origin of, their properties and characteristics that distinguish from other cells, the classification, and biological disorders causing neurodegenerative diseases. The literature, data and arguments that are dealing with, how the differentiate into neural cells, and how could this process can be handled in vitro are reviewed. The unique capability of these cells to form various tissues under definite signals received from the body, it makes this cell an object of extensive research. Subsequently, information has been compiled on the question how neural differentiation is controlled on the molecular level, and controlled in vivo. Finally one major gene involved will be investigated, which are may be investigated in practical pirogue according there expression patterns. And this gene is tyrosine hydroxylase.

Human Cardiomyocytes Prior to Birth by Integration-Free Reprogramming of Amniotic Fluid Cells

The establishment of an abundant source of autologous cardiac progenitor cells would represent a major advance toward eventual clinical translation of regenerative medicine strategies in children with prenatally diagnosed congenital heart disease. In support of this concept, we sought to examine whether functional, transgene‐free human cardiomyocytes (CMs) with potential for patient‐specific and autologous applications could be reliably generated following routine amniocentesis. Under institutional review board approval, amniotic fluid specimens (8–10 ml) at 20 weeks gestation were expanded and reprogrammed toward pluripotency using nonintegrating Sendai virus (SeV) expressing OCT4, SOX2, cMYC, and KLF4. Following exposure of these induced pluripotent stem cells to cardiogenic differentiation conditions, spontaneously beating amniotic fluid‐derived cardiomyocytes (AF‐CMs) were successfully generated with high efficiency. After 6 weeks, quantitative gene expression revealed a mixed population of differentiated atrial, ventricular, and nodal AF‐CMs, as demonstrated by upregulation of multiple cardiac markers, including MYH6, MYL7, TNNT2, TTN, and HCN4, which were comparable to levels expressed by neonatal dermal fibroblast‐derived CM controls. AF‐CMs had a normal karyotype and demonstrated loss of NANOG, OCT4, and the SeV transgene. Functional characterization of SIRPA+ AF‐CMs showed a higher spontaneous beat frequency in comparison with dermal fibroblast controls but revealed normal calcium transients and appropriate chronotropic responses after β‐adrenergic agonist stimulation. Taken together, these data suggest that somatic cells present within human amniotic fluid can be used to generate a highly scalable source of functional, transgene‐free, autologous CMs before a child is born. This approach may be ideally suited for patients with prenatally diagnosed cardiac anomalies.

Cohort Profile: The Heinz C. Prechter Longitudinal Study of Bipolar Disorder

Cohort Profile: The Heinz C. Prechter Longitudinal Study of Bipolar Disorder Melvin G McInnis,* Shervin Assari, Masoud Kamali, Kelly Ryan, Scott A Langenecker, Erika FH Saunders, Kritika Versha, Simon Evans, K Sue O’Shea, Emily Mower Provost, David Marshall, Daniel Forger, Patricia Deldin and Sebastian Zoellner; for the Prechter Bipolar Clinical Research Collaborative Department of Psychiatry, University of Michigan, Ann Arbor, MI, USA, Department of Psychiatry, Massachusetts General Hospital, Boston, MA, USA, Department of Psychiatry, University of Illinois at Chicago, Chicago, IL, USA, Department of Psychiatry, Penn State Hershey Medical Group, Hershey, PA, USA, Department of Cell and Developmental Biology, Department of Computer Science and Engineering, Department of Mathematics, Department of Psychology and Department of Biostatistics, University of Michigan, Ann Arbor, MI, USA

An apicosome initiates self-organizing morphogenesis of human pluripotent stem cells

Human pluripotent stem cells (hPSCs) self-organize into apicobasally polarized cysts, reminiscent of the lumenal epiblast stage, providing a model to explore key morphogenic processes in early human embryos. Here, we show that apical polarization begins on the interior of single hPSCs through the dynamic formation of a highly organized perinuclear apicosome structure. The membrane surrounding the apicosome is enriched in apical markers and displays microvilli and a primary cilium; its lumenal space is rich in Ca2+. Time-lapse imaging of isolated hPSCs reveals that the apicosome forms de novo in interphase, retains its structure during mitosis, is asymmetrically inherited after mitosis, and relocates to the recently formed cytokinetic plane, where it establishes a fully polarized lumen. In a multicellular aggregate of hPSCs, intracellular apicosomes from multiple cells are trafficked to generate a common lumenal cavity. Thus, the apicosome is a unique preassembled apical structure that can be rapidly used in single or clustered hPSCs to initiate self-organized apical polarization and lumenogenesis.

Use of the Sleeping Beauty transposon system for stable gene expression in mouse embryonic stem cells.

Sleeping Beauty (SB) transposon-based transfection is a two-component system consisting of a transposase and a transposon containing inverted repeat/direct repeat (IR/DR) sequences that result in precise integration into a TA dinucleotide. The transposon is designed with an expression cassette of interest flanked by IR/DRs, and SB transposase mediates stable integration and reliable long-term expression of the gene of interest. It has recently been demonstrated that SB efficiently mediates gene transfer and stable gene expression in human embryonic stem (ES) cells. Here, we describe a method for transfecting and establishing stable cell lines in mouse embryonic stem (mES) cells with the SB system.