Pre-clinical evaluation of clinically relevant iPS cell derived neuroepithelial stem cells as an off-the-shelf cell therapy for spinal cord injury.

Preclinical transplantations using human neuroepithelial stem (NES) cells in spinal cord injury models have exhibited promising results and demonstrated cell integration and functional improvement in transplanted animals. Previous studies have relied on the generation of research grade cell lines in continuous culture. Using fresh cells presents logistic hurdles for clinical transition regarding time and resources for maintaining high quality standards.

Age-related pathological impairments in directly reprogrammed dopaminergic neurons derived from patients with idiopathic Parkinson's disease.

We have developed an efficient approach to generate functional induced dopaminergic (DA) neurons from adult human dermal fibroblasts. When performing DA neuronal conversion of patient fibroblasts with idiopathic Parkinson's disease (PD), we could specifically detect disease-relevant pathology in these cells. We show that the patient-derived neurons maintain age-related properties of the donor and exhibit lower basal chaperone-mediated autophagy compared with healthy donors.

Presynaptic dysfunction in CASK-related neurodevelopmental disorders.

CASK-related disorders are genetically defined neurodevelopmental syndromes. There is limited information about the effects of CASK mutations in human neurons. Therefore, we sought to delineate CASK-mutation consequences and neuronal effects using induced pluripotent stem cell-derived neurons from two mutation carriers.

DNA methylation changes in Down syndrome derived neural iPSCs uncover co-dysregulation of ZNF and HOX3 families of transcription factors.

Background: Down syndrome (DS) is characterized by neurodevelopmental abnormalities caused by partial or complete trisomy of human chromosome 21 (T21). Analysis of Down syndrome brain specimens has shown global epigenetic and transcriptional changes but their interplay during early neurogenesis remains largely unknown. We differentiated induced pluripotent stem cells (iPSCs) established from two DS patients with complete T21 and matched euploid donors into two distinct neural stages corresponding to early- and mid-gestational ages.

Single cell analysis of autism patient with bi-allelic NRXN1-alpha deletion reveals skewed fate choice in neural progenitors and impaired neuronal functionality.

We generated human iPS derived neural stem cells and differentiated cells from healthy control individuals and an individual with autism spectrum disorder carrying bi-allelic NRXN1-alpha deletion. We investigated the expression of NRXN1-alpha during neural induction and neural differentiation and observed a pivotal role for NRXN1-alpha during early neural induction and neuronal differentiation. Single cell RNA-seq pinpointed neural stem cells carrying NRXN1-alpha deletion shifting towards radial glia-like cell identity and revealed higher proportion of differentiated astroglia.

Overactive BRCA1 Affects Presenilin 1 in Induced Pluripotent Stem Cell-Derived Neurons in Alzheimer's Disease.

The BRCA1 protein, one of the major players responsible for DNA damage response has recently been linked to Alzheimer's disease (AD). Using primary fibroblasts and neurons reprogrammed from induced pluripotent stem cells (iPSC) derived from familial AD (FAD) patients, we studied the role of the BRCA1 protein underlying molecular neurodegeneration. By whole-transcriptome approach, we have found wide range of disturbances in cell cycle and DNA damage response in FAD fibroblasts.

Integration Free Derivation of Human Induced Pluripotent Stem Cells Using Laminin 521 Matrix.

Xeno-free and fully defined conditions are key parameters for robust and reproducible generation of homogenous human induced pluripotent stem (hiPS) cells. Maintenance of hiPS cells on feeder cells or undefined matrices are susceptible to batch variances, pathogenic contamination and risk of immunogenicity. Utilizing the defined recombinant human laminin 521 (LN-521) matrix in combination with xeno-free and defined media formulations reduces variability and allows for the consistent generation of hiPS cells.

Derivation of human iPS cell lines from monozygotic twins in defined and xeno free conditions.

Human induced pluripotent stem (hiPS) cell lines CTRL-9-II and CTRL-10-I were derived from healthy monozygotic twin donors using non-integrating RNA based Sendai virus reprogramming and cultured in a xeno-free chemically defined condition. The established hiPS cell lines, CTRL-9-II and CTRL-10-I, are karyotypically normal, free from reprogramming vectors, display endogenously expression of pluripotency factors at levels similar to embryonic stem cells. The generated iPS cell lines demonstrate pluripotency by passing bioinformatics assay PluriTest and by embryonic body assay.

Generation of human iPS cell line CTL07-II from human fibroblasts, under defined and xeno-free conditions.

CTL07-II is a healthy feeder-free and characterized human induced pluripotent stem (iPS) cell line. Cultured under xeno-free and defined conditions. The line is generated from healthy human fibroblasts with non-integrating Sendai virus vectors encoding the four Yamanaka factors, OCT4, SOX2, KLF4 and cMYC.

Performance of idealized column structures under high pressure.

We have examined the influence of the longitudinal temperature and pressure gradients in columns operated under very high pressures on the coefficients of the van Deemter equation under the idealized condition of complete radial uniformity. These gradients change the diffusion coefficients over the length of the column, and the equation takes a new form, where the classical linear C-term is replaced by more complex forms that capture the effects of these axial gradients. The details of the derivations are shown and the implications are discussed.

Implications of column peak capacity on the separation of complex peptide mixtures in single- and two-dimensional high-performance liquid chromatography.

Column peak capacity was utilized as a measure of column efficiency for gradient elution conditions. Peak capacity was evaluated experimentally for reversed-phase (RP) and cation-exchange high-performance liquid chromatography (HPLC) columns, and compared to the values predicted from RP-HPLC gradient theory. The model was found to be useful for the prediction of peak capacity and productivity in single- and two-dimensional (2D) chromatography.

Moment analysis of mass-transfer kinetics in C18-silica monolithic columns.

The moment analysis of elution peak profiles based on new moment equations provides information on the mass-transfer characteristics of C(18)-silica monolithic columns. The flow rate dependence of the HETP data was analyzed using the generalized van Deemter equation, after correction of these data by subtraction of the external mass-transfer contribution to band broadening. Kinetic parameters and diffusion coefficients related to the mass-transfer processes in monolithic columns were derived by taking advantage of the different flow velocity dependence of their contributions to band broadening.

Influence of the pH on the behavior of an imprinted polymeric stationary phase--supporting evidence for a binding site model.

The equilibrium isotherms of the two enantiomers of phenylalanine anilide (PA) were measured by conventional frontal analysis at three different pH on a thermally-treated imprinted stationary phase selective for the L enantiomer. The first of these pH (buffer pH=3.0, pH(app)=4.

Repeatability and reproducibility of retention data and band profiles on reversed-phase liquid chromatography columns. V. Results obtained with Vydac 218TP C18 columns.

The general goal of this work is to investigate the precision of chromatographic data and to determine which properties of chromatographic columns influence this factor. Chromatographic data were acquired under five different sets of experimental conditions for 30 neutral, acidic and basic test compounds on columns packed with Vydac 218TP C18, a polymeric, wide-pore silica-based stationary phase. Five columns packed with samples from the same batch of this packing material were used to measure the column-to-column reproducibility and six columns packed with material from six different batches to measure the batch-to-batch reproducibility.

Identification of the factors that influence the reproducibility of chromatographic retention data.

Principal component analysis was used to identify the parameters that influence the column-to-column and batch-to-batch reproducibility of retention times and retention factors measured on Symmetry C18, Kromasil C18, Luna C18 (2) and Vydac RP C18, all reversed-phase silica columns. We devised a procedure that allows the determination of the differences in column volume and packing density between two columns, provided that these columns are packed with identical stationary phases (i.e.

Comparison of the repeatability of quantitative data measured in high-performance liquid chromatography with UV and atmospheric pressure chemical ionization mass spectrometric detection.

A detailed comparison of the repeatability of the retention times, the peak efficiencies and the peak areas of a dozen probe compounds achieved in HPLC, using either HPLC-UV or HPLC-MS for detection purpose, is reported. Three groups of conventional analytes, each one separated under a different set of experimental conditions, were selected for this study. Most of the compounds are basic, the other ones being neutral.

Repeatability and reproducibility of retention data and band profiles on reversed-phase liquid chromatography columns. IV. Results obtained with Luna C18 (2) columns.

The reproducibility of retention data and band profile characteristics was investigated for a series of columns packed with Luna C18 (2), a silica-based, reversed-phase adsorbent. High precision data were obtained and statistically compared among five columns from the same batch (column-to-column reproducibility) and nine columns from as many different batches (batch-to-batch reproducibility). These data were acquired under five different sets of chromatographic conditions, for a group of 30 neutral, acidic, and basic compounds selected as probes following an experimental protocol previously described.

Repeatability and reproducibility of retention data and band profiles on reversed-phase liquid chromatography columns. III. Results obtained with Kromasil C18 columns.

The reproducibility of the retention data and the band profiles was investigated with Kromasil C18 columns (silica-based monomeric type reversed-phase packing material). High precision data were obtained and statistically compared among five columns from the same batch (column-to-column reproducibility) and six columns, one from each of six different batches (batch-to-batch reproducibility). These data were acquired under five different sets of chromatographic conditions, for a group of 30 neutral, acidic and basic compounds selected as probes following an experimental protocol previously described.

Influence of Thermal Annealing on the Thermodynamic and Mass-Transfer Kinetic Properties of d- and l-Phenylalanine Anilide on Imprinted Polymeric Stationary Phases.

An investigation of the material, chromatographic, thermodynamic, and kinetic properties of thermally treated (i.e., annealed) polymeric stationary phases imprinted with l-phenylalanine anilide (l-PA) was carried out.

Determination of serotonin released from coffee wax by liquid chromatography.

A simple hydrolysis and extraction method was developed for the release of serotonin (5-hydroxytryptamine) from a coffee wax sample obtained from decaffeination of coffee beans. The recoverable amount of serotonin was determined by reversed-phase high-performance liquid chromatography with gradient elution and UV detection, using the standard addition method. Different type of basic deactivated chromatographic columns were used for the separation.