Gaussian mixture models and machine learning predict megakaryocytic growth and differentiation potential ex vivo

The ability to analyze single cells via flow cytometry has resulted in a wide range of biological and medical applications. Currently, there is no established framework to compare and interpret time-series flow cytometry data for cell engineering applications. Manual analysis of temporal trends is time-consuming and subjective for large-scale datasets. We resolved this bottleneck by developing TEmporal Gaussian Mixture models (TEGM), an unbiased computational strategy to quantify and predict temporal trends of developing cell subpopulations indicative of cellular phenotype. TEGM applies Gaussian mixture models and gradient boosted trees for cell engineering applications. TEGM enables the extraction of subtle features, such as the dispersion and rate of change of surface marker expression for each subpopulation over time. These critical, yet hard-to-discern, features are fed into machine-learning algorithms that predict underlying cell classes. Our framework can be flexibly applied to conventional flow cytometry sampling schemes, and allows for faster and more consistent processing of time-series flow cytometry data. Please click Additional Files below to see the full abstract

Using Gaussian mixture models and machine learning to predict donor- dependent megakaryocytic cell growth and differentiation potential ex vivo

The ability to analyze single cells via flow cytometry has resulted in a wide range of biological and medical applications. Currently, there is no established framework to compare and interpret time-series flow cytometry data for cell engineering applications. Manual analysis of temporal trends is time-consuming and subjective for large-scale datasets. We resolved this bottleneck by developing TEmporal Gaussian Mixture models (TEGM), an unbiased computational strategy to quantify and predict temporal trends of developing cell subpopulations indicative of cellular phenotype.. Please click Additional Files below to see the full abstract

N′-[(E)-1-(4-Chloro­phen­yl)ethyl­idene]-2-[4-(2-methyl­prop­yl)phen­yl]propano­hydrazide

The asymmetric unit of the title compound, C21H25ClN2O, contains four crystallographically independent mol­ecules, which differ mainly in the orientation of the isobutyl groups. The benzene rings are almost orthogonal to each other, forming dihedral angles of 87.40 (6), 88.69 (6), 84.88 (6) and 85.12 (6)° in the four mol­ecules. The crystal structure is stabilized by inter­molecular N—H⋯O and C—H⋯O hydrogen bonds, together with C—H⋯π inter­actions

2-(4-Isobutyl­phen­yl)-N′-[1-(4-nitro­phen­yl)ethyl­idene]propanohydrazide

The mol­ecule of the title compound, C21H25N3O3, exists in a trans configuration with respect to the ethyl­idene unit. The dihedral angle between the two substituted benzene rings is 86.99 (7)°. The nitro group is twisted from the attached benzene ring at an angle of 17.02 (7)°. In the crystal structure, mol­ecules are linked by pairs of N—H⋯O hydrogen bonds in a face-to-face manner into centrosymmetric dimers. These dimer units are further linked into chains along the c axis by weak C—H⋯O inter­actions. These chains are stacked along the b axis. The crystal is further stabilized by weak C—H⋯π inter­actions

N′-[(E)-4-Bromo­benzyl­idene]-2-(4-isobutyl­phen­yl)propanohydrazide

The asymmetric unit of the title compound, C20H23BrN2O, contains two independent mol­ecules (A and B), in which the orientations of the 4-isobutyl­phenyl units are different. The dihedral angle between the two benzene rings is 88.45 (8)° in mol­ecule A and 89.87 (8)° in mol­ecule B. Mol­ecules A and B are linked by a C—H⋯N hydrogen bond. In the crystal, mol­ecules are linked into chains running along the a axis by inter­molcular N—H⋯O and C—H⋯O hydrogen bonds. The crystal structure is further stabilized by C—H⋯π inter­actions. The presence of pseudosymmetry in the structure suggests the higher symmetry space group Pbca. However, attempts to refine the structure in this space group resulted in a disorder model with high R (0.097) and wR (0.257) values. The crystal studied was an inversion twin with a 0.595 (4):0.405 (4) domain ratio

2-[4-(2-Methyl­prop­yl)phen­yl]-N′-[(E)-1-phenyl­ethyl­idene]propane­hydrazide

In the title compound, C21H26N2O, the dihedral angle between the two aromatic rings is 85.90 (19)°. The propenone–hydrazide unit forms dihedral angles of 21.62 (8) and 72.83 (9)°, respectively, with the terminal and central aromatic rings. The 2-methyl­propyl group is disordered over two sites, with occupancies of 0.533 (13) and 0.467 (13). In crystal structure, mol­ecules are linked into centrosymmetric dimers by paired N—H⋯O and C—H⋯O hydrogen bonds. In addition, C—H⋯π inter­actions are observed

A new polymorph of N-(2-{N′-[(1E)-2-hy­dr­oxy­benzyl­­idene]hydrazinecarbon­yl}phen­yl)benzamide

The title compound, C 21 H 17 N 3 O 3 , is a new polymorph of an already published structure [Shashidhar et al. (2006). Acta Cryst. E62, o4473-o4475]. The previously reported structure crystallizes in the monoclinic space group C2/c, whereas the structure reported here is in the tetragonal space group I4 1 /a. The bond lengths and angles are similar in both structures. The molecule adopts an extended conformation via intramolecular N-H⋯O and O-H⋯N hydrogen bonds; the terminal phenyl ring and the hydroxylphenyl ring are twisted with respect to the central benzene ring by 44.43 (7) and 21.99 (8)°, respectively. In the crystal, molecules are linked by N-H⋯O hydrogen bonds, weak C-H⋯O hydrogen bonds and weak C-H⋯π interactions into a three-dimensional supramolecular network

TRAIL Death Receptor-4, Decoy Receptor-1 and Decoy Receptor-2 Expression on CD8+ T Cells Correlate with the Disease Severity in Patients with Rheumatoid Arthritis

BACKGROUND: Rheumatoid Arthritis (RA) is a chronic autoimmune inflammatory disorder. Although the pathogenesis of disease is unclear, it is well known that T cells play a major role in both development and perpetuation of RA through activating macrophages and B cells. Since the lack of TNF-Related Apoptosis Inducing Ligand (TRAIL) expression resulted in defective thymocyte apoptosis leading to an autoimmune disease, we explored evidence for alterations in TRAIL/TRAIL receptor expression on peripheral T lymphocytes in the molecular mechanism of RA development. METHODS: The expression of TRAIL/TRAIL receptors on T cells in 20 RA patients and 12 control individuals were analyzed using flow cytometry. The correlation of TRAIL and its receptor expression profile was compared with clinical RA parameters (RA activity scored as per DAS28) using Spearman Rho Analysis. RESULTS: While no change was detected in the ratio of CD4+ to CD8+ T cells between controls and RA patient groups, upregulation of TRAIL and its receptors (both death and decoy) was detected on both CD4+ and CD8+ T cells in RA patients compared to control individuals. Death Receptor-4 (DR4) and the decoy receptors DcR1 and DcR2 on CD8+ T cells, but not on CD4+ T cells, were positively correlated with patients' DAS scores. CONCLUSIONS: Our data suggest that TRAIL/TRAIL receptor expression profiles on T cells might be important in revelation of RA pathogenesis

Ret is essential to mediate GDNF’s neuroprotective and neuroregenerative effect in a Parkinson disease mouse model

Glial cell line-derived neurotrophic factor (GDNF) is a potent survival and regeneration-promoting factor for dopaminergic neurons in cell and animal models of Parkinson disease (PD). GDNF is currently tested in clinical trials on PD patients with so far inconclusive results. The receptor tyrosine kinase Ret is the canonical GDNF receptor, but several alternative GDNF receptors have been proposed, raising the question of which signaling receptor mediates here the beneficial GDNF effects. To address this question we overexpressed GDNF in the striatum of mice deficient for Ret in dopaminergic neurons and subsequently challenged these mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Strikingly, in this established PD mouse model, the absence of Ret completely abolished GDNF’s neuroprotective and regenerative effect on the midbrain dopaminergic system. This establishes Ret signaling as absolutely required for GDNF’s effects to prevent and compensate dopaminergic system degeneration and suggests Ret activation as the primary target of GDNF therapy in PD

NRF2 Activation Restores Disease Related Metabolic Deficiencies in Olfactory Neurosphere-Derived Cells from Patients with Sporadic Parkinson's Disease

Extent: 14p.Background: Without appropriate cellular models the etiology of idiopathic Parkinson’s disease remains unknown. We recently reported a novel patient-derived cellular model generated from biopsies of the olfactory mucosa (termed olfactory neurosphere-derived (hONS) cells) which express functional and genetic differences in a disease-specific manner. Transcriptomic analysis of Patient and Control hONS cells identified the NRF2 transcription factor signalling pathway as the most differentially expressed in Parkinson’s disease. Results: We tested the robustness of our initial findings by including additional cell lines and confirmed that hONS cells from Patients had 20% reductions in reduced glutathione levels and MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)- 2-(4-sulfophenyl)-2H-tetrazolium, inner salt] metabolism compared to cultures from healthy Control donors. We also confirmed that Patient hONS cells are in a state of oxidative stress due to higher production of H2O2 than Control cultures. siRNA-mediated ablation of NRF2 in Control donor cells decreased both total glutathione content and MTS metabolism to levels detected in cells from Parkinson’s Disease patients. Conversely, and more importantly, we showed that activation of the NRF2 pathway in Parkinson’s disease hONS cultures restored glutathione levels and MTS metabolism to Control levels. Paradoxically, transcriptomic analysis after NRF2 pathway activation revealed an increased number of differentially expressed mRNAs within the NRF2 pathway in L-SUL treated Patient-derived hONS cells compared to L-SUL treated Controls, even though their metabolism was restored to normal. We also identified differential expression of the PI3K/AKT signalling pathway, but only post-treatment. Conclusions: Our results confirmed NRF2 as a potential therapeutic target for Parkinson’s disease and provided the first demonstration that NRF2 function was inducible in Patient-derived cells from donors with uniquely varied genetic backgrounds. However, our results also demonstrated that the response of PD patient-derived cells was not co-ordinated in the same way as in Control cells. This may be an important factor when developing new therapeutics.Anthony L. Cook, Alejandra M. Vitale, Sugandha Ravishankar, Nicholas Matigian, Greg T. Sutherland, Jiangou Shan, Ratneswary Sutharsan, Chris Perry, Peter A. Silburn, George D. Mellick, Murray L. Whitelaw, Christine A. Wells, Alan Mackay-Sim and Stephen A. Woo