Homologous feeder cells support undifferentiated growth and pluripotency in monkey embryonic stem cells

In the present study, five homologous feeder cell lines were developed for the culture and maintenance of rhesus monkey embryonic stem cells (rESCs). Monkey ear skin fibroblasts (MESFs), monkey oviductal fibroblasts (MOFs), monkey follicular granulosa fibroblast-like (MFG) cells, monkey follicular granulosa epithelium-like (MFGE) cells, and clonally derived fibroblasts from MESF (CMESFs) were established and compared with the ability of mouse embryonic fibroblasts (MEFs) to support rESC growth. MESF, MOF, MFG, and CMESF cells, but not MFGE cells, were as good as or better than MEFs in supporting undifferentiated growth while maintaining the differentiation potential of the rESCs. In an effort to understand the unique properties of supportive feeder cells, expression levels for a number of candidate genes were examined. MOF, MESF, and MEF cells highly expressed leukemia inhibitory factor, ciliary neurotrophic factor, basic fibroblast growth factor, stem cell factor, transforming growth factor PI, bone morphogenetic protein 4, and WNT3A, whereas WNT2, WNT4, and WNT5A were downregulated, compared with MFGE cells. Additionally, all monkey feeder cell lines expressed Dkk1 and LRP6, antagonists of the WNT signaling pathway, but not WNT1, WNT8B, or Dkk2. rESCs grown on homologous feeders maintained normal karyotypes, displayed the characteristics of ESCs, including morphology, alkaline phosphatase, Oct4, the cell surface markers stage-specific embryonic antigen (SSEA)-3, SSEA-4, tumor-related antigen (TRA)-1-60, and TRA-1-81, and formed cystic embryoid bodies in vitro that included differentiated cells representing the three major germ layers. These results indicate that the four homologous feeder cell lines can be used to support the undifferentiated growth and maintenance of pluripotency in rESCs.

Hepatocyte growth factor enhances the generation of high-purity oligodendrocytes from human embryonic stem cells

Generation of homogeneous oligodendrocytes as donor cells is essential for human embryonic stem cell (hESC)-based cell therapy for demylinating diseases. Herein we present a novel method for efficiently obtaining mature oligodendrocytes from hESCs with high purity (79.7 +/- 6.9%), using hepatocyte growth factor (HGF) and G5 supplement(containing insulin, transferrin, selenite, biotin, hydrocortisone, basic fibroblast growth factor and epidermal growth factor) in a four-step method. We induced hESCs into neural progenitors (NP) with HGF (5 ng/ml) and G5 (1 x) supplemented medium in an adherent differentiation system. The purified NPs were amplified in suspension as neurospheres for 1 month, and terminal oligodendrocyte differentiation was then induced by G5 supplement withdrawal and HGF treatment (20 ng/ml). The cells generated displayed typical morphologies of mature oligodendrocytes and expressed oligodendrocyte markers O4 and myelin basic protein (MBP). Our result revealed that HGF significantly enhanced the proliferation of hESC-derived NPs and promoted the differentiation as well as the maturation of oligodendrocytes from NPs. Further studies suggest that HGF/c-Met signaling pathway might play an important role in oligodendrocyte differentiation in our system. Our studies provide a means for generating the clinically relevant cell type and a platform for deciphering the molecular mechanisms that control oligodendrocyte differentiation. (C) 2009 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved.

Growth of GaSb and GaInAsSb layers for thermophotovolatic cells by liquid phase epitaxy - art. no. 68411E

GaSb based cells as receivers in thermophotovoltaic system have attracted great interest and been extensively studied in the recent 15 years. Although nowadays the manufacturing technologies have made a great progress, there are still some details need to make a further study. In this paper, undoped and doped GaSb layers were grown on n-GaSb (100) substrates from both Ga-rich and Sb-rich solutions using liquid phase epitaxy (LPE) technique. The nominal segregation coefficients k of intentional doped Zn were 1.4 and 8.8 determined from the two kinds of GaSb epitaxial layers. Additionally, compared with growing from Ga-rich solutions, the growing processes from Sb-rich solutions were much easier to control and the surface morphologies of epitaxial layers were smoother. Further-more, in order to broaden the absorbing edge, Ga1-xInxAsySb1-y quaternary alloys were grown on both GaSb and InAs substrates from In-rich solutions, under different temperature respectively.

Electroactive Aniline Pentamer Cross-Linking Chitosan for Stimulation Growth of Electrically Sensitive Cells

A new kind of electroactive polymers was synthesized by using aniline pentamer (AP) cross-linking chitosan (CS) in acetic acid/DMSO/DMF solution. UV-vis and CV confirmed the electroactivity of polymers in acidic aqueous solution. The amphiphilic polymers self-assembled into 200-300 nm micelles by dialysis against deionized water from the acetic acid buffer solution. Three samples with different weight percentages of AP were used to identify the relationship between the content of AP and the differentiation of rat neuronal pheochromocytoma PC-12 cells without external stimulation.

Telomerase Activity and Microgravity-Dependent Apoptosis in Human Leukemia HL-60 Cells

Mammalian cells subjected to conditions of spaceflight and the microgravity environment ofspace; manifest a number of alterations in structure and function. Among the most notable changes incells flown on the Space Shuttle are reduced growth activation and decline in growth rate in the totalpopulation. Other changes include chromosomal aberrations, inhibited locomotion, alteredcytokine production, changes in PKC distribution, and increased apoptos.

A Novel Counter Sheet-Flow Sandwich Cell Culture Device For Mammalian Cell Growth In Space

Cell culture and growth in space is crucial to understand the cellular responses under microgravity. The effects of microgravity were coupled with such environment restrictions as medium perfusion, in which the underlying mechanism has been poorly understood. In the present work, a customer-made counter sheet-flow sandwich cell culture device was developed upon a biomechanical concept from fish gill breathing. The sandwich culture unit consists of two side chambers where the medium flow is counter-directional, a central chamber where the cells are cultured, and two porous polycarbonate membranes between side and central chambers. Flow dynamics analysis revealed the symmetrical velocity profile and uniform low shear rate distribution of flowing medium inside the central culture chamber, which promotes sufficient mass transport and nutrient supply for mammalian cell growth. An on-orbit experiment performed on a recovery satellite was used to validate the availability of the device.

Transplantable neural progenitor populations derived from Rhesus monkey embryonic stem cells

Cell-based therapies using embryonic stem cells (ESCs) in the treatment of neural disease will require the generation of homogenous donor neural progenitor (NP) populations. Here we describe an efficient culture system containing hepatocyte growth factor (HGF) and G5 supplement for the production of highly enriched (88.3% +/- 8.1%)populations of NPs from rhesus monkey ESCs. Additional purification resulted in NP preparations that were 98% nestin positive. Moreover, NPs, as monolayers or neurospheres, could be maintained for prolonged periods of time in media containing HGF+G5 or G5 alone. In vitro differentiation and in vivo transplantation assays showed that NPs could differentiate into neurons, astrocytes, and oligodendrocytes. The kinds and quantities of differentiated cells derived from NPs were closely correlated with their niches in vivo. Glial differentiation was predominant in periventricular areas, whereas cells migrating into the cortex were mostly neurons. Cell counts showed that 2 months after transplantation, approximately 25% of transplanted NPs survived and 65% - 80% of the surviving transplanted cells migrated along the ventricular wall or in a radial fashion. Subcloning demonstrated that several clonal lines derived from NPs expressed nestin and differentiated into three neural lineages in vitro and in rat brains in vivo. In contrast, some subcloned lines showed restricted differentiation both in vitro and in vivo in rat brains. These observations set the stage for obtaining highly enriched NPs and evaluating the efficacy of NP-based transplantation therapy in the nonhuman primate and will provide a platform for probing the molecular mechanisms that control neural induction.

Dissecting Signaling Pathways That Govern Self-renewal of Rabbit Embryonic Stem Cells

The pluripotency and self-renewal of embryonic stem cells (ESC) are regulated by a variety of cytokines/growth factors with some species differences. We reported previously that rabbit ESC (rESC) are more similar to primate ESC than to mouse ESC. However,

Neural progenitors derived from monkey embryonic stem cells in a simple monoculture system

A simple monoculture system. combined with a chemically defined medium containing hepatocyte growth factor (HGF) and G5 supplement, was used to induce rhesus monkey embryonic stem cells (rESC) directly into neuroepithelial (NE) cells. Under these conditio

Expression of VEGFR-2 on HaCaT cells is regulated by VEGF and plays an active role in mediating VEGF induced effects

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 play important roles in mitogenesis and chemotaxis of endothelial cells. In normal human skin, VEGF is expressed and secreted by epidermal keratinocytes. Emerging data suggest that keratin