Neural differentiation from human embryonal carcinoma stem cells

It is understood that retinoic acid (RA), sonic hedgehog (Shh) and bone morphogenic proteins (BMPs) play an important role in cell fate determination and the specification of inter-neurons and motor neurons along the dorsal-ventral axis in the neural tube. In this study, we investigated the function of these signalling molecules to instruct the differentiation of human pluripotent stem cells to form specific neuronal subtypes. TERA2.cl.SP12 embryonal carcinoma (EC) cells are a robust caricature of human embryogenesis and an accepted model of neural differentiation. Gene and protein expression analyses using RT-PCR, western blotting and immunocytochemical techniques indicated that human EC cells respond to RA, BMPs and Shh in a conserved manner and regulate neural transcription factors and structural proteins in a predicted way as cells commit toward the motor neuron phenotype. To assess the function of these differentiated neurons, we tested their ability to innervate skeletal muscle myotubes and induce muscle cell contraction. Myotubes contracted only when cocultured with neurons. The number of contractile events increased significantly when cells differentiated into motor neurons were cocultured with myotubes compared to cocultures with cells that formed intemeurons. Staining for α-bungarotoxin showed positive staining in a pattern characteristic of boutons found in neuromuscular junctions. We also showed that muscle contraction could be manipulated pharmacologically: curare and atropine blocked myotube contraction, whereas acetylcholine and carbachol increased the number of contractile events. In other experiments, we have also shown that cells exposed to RA and Shh in conjunction with other growth factors over different time periods, preferentially form oligodendrocytes and/or interneurons. These results indicate it is feasible to control and direct the differentiation of human stem cells and produce specific neuron subtypes in vitro. Furthermore, this system acts as a useful model to investigate the molecular mechanisms and signalling pathways that control neural differentiation in man

Realization of multiple charge density waves in NbTe2 at the monolayer limit

Abstract: Layered transition-metal dichalcogenides (TMDCs) down to the monolayer (ML) limit provide a fertile platform for exploring charge-density waves (CDWs). Though bulk NbTe2 is known to harbor a single axis 3*1 CDW coexisting with non-trivial quantum properties, the scenario in the ML limit is still experimentally unknown. In this study, we unveil the richness of the CDW phases in ML NbTe2, where not only the theoretically predicted 4*4 and 4*1 phases, but also two unexpected sqrt(28)*sqrt(28) and sqrt(19)*sqrt(19) phases, can be realized. For such a complex CDW system, we establish an exhaustive growth phase diagram via systematic efforts in the material synthesis and scanning tunneling microscope characterization. Moreover, we report that the energetically stable phase is the larger scale order (sqrt(19)*sqrt(19)), which is surprisingly in contradiction to the prior prediction (4*4). These findings are confirmed using two different kinetic pathways, i.e., direct growth at proper growth temperatures (T), and low-T growth followed by high-T annealing. Our results provide a comprehensive diagram of the "zoo" of CDW orders in ML 1T-NbTe2 for the first time and offer a new material platform for studying novel quantum phases in the 2D limit

SARS-CoV-2 infection causes dopaminergic neuron senescence

COVID-19 patients commonly present with signs of central nervous system and/or peripheral nervous system dysfunction. Here, we show that midbrain dopamine (DA) neurons derived from human pluripotent stem cells (hPSCs) are selectively susceptible and permissive to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. SARS-CoV-2 infection of DA neurons triggers an inflammatory and cellular senescence response. High-throughput screening in hPSC-derived DA neurons identified several FDA-approved drugs that can rescue the cellular senescence phenotype by preventing SARS-CoV-2 infection. We also identified the inflammatory and cellular senescence signature and low levels of SARS-CoV-2 transcripts in human substantia nigra tissue of COVID-19 patients. Furthermore, we observed reduced numbers of neuromelanin+ and tyrosine-hydroxylase (TH)+ DA neurons and fibers in a cohort of severe COVID-19 patients. Our findings demonstrate that hPSC-derived DA neurons are susceptible to SARS-CoV-2, identify candidate neuroprotective drugs for COVID-19 patients, and suggest the need for careful, long-term monitoring of neurological problems in COVID-19 patients.</p

SARS-CoV-2 infection causes dopaminergic neuron senescence

COVID-19 patients commonly present with signs of central nervous system and/or peripheral nervous system dysfunction. Here, we show that midbrain dopamine (DA) neurons derived from human pluripotent stem cells (hPSCs) are selectively susceptible and permissive to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. SARS-CoV-2 infection of DA neurons triggers an inflammatory and cellular senescence response. High-throughput screening in hPSC-derived DA neurons identified several FDA-approved drugs that can rescue the cellular senescence phenotype by preventing SARS-CoV-2 infection. We also identified the inflammatory and cellular senescence signature and low levels of SARS-CoV-2 transcripts in human substantia nigra tissue of COVID-19 patients. Furthermore, we observed reduced numbers of neuromelanin+ and tyrosine-hydroxylase (TH)+ DA neurons and fibers in a cohort of severe COVID-19 patients. Our findings demonstrate that hPSC-derived DA neurons are susceptible to SARS-CoV-2, identify candidate neuroprotective drugs for COVID-19 patients, and suggest the need for careful, long-term monitoring of neurological problems in COVID-19 patients.</p

Abstract 2333: The atypical tumor suppressor p27 regulates cellular proliferation, invasion, and metastasis via subcellular localization in distinct microenvironments

Abstract While mutational loss of the p27 cell cycle regulatory protein is rare, oncogenic deregulation of p27 via accelerated degradation or mislocalization to the cytoplasm is commonly observed in human tumors. A growing body of evidence indicates that, when mislocalized to the cytoplasm, p27 promotes cell motility via RhoA-ROCK signaling inhibition. The present data explores the role of p27 in the regulation of both orthotopic mammary tumor growth and ectopic pulmonary metastasis in the MDA-MB-231-4175 model of malignant human breast cancer. The MDA-MB-231-4175 cell line (hereafter 4175) shows much greater metastasis to the lungs in murine xenograft models. Relative to the parental MDA-MB-231 cells, the metastatic 4175 derivative demonstrates activation of PI3K/mTOR signaling. These cells have elevated pmTOR, pAKTS473, pSGK1, and pRSK1. This results in increased total p27 and, when equal levels of total p27 are titrated, the highly metastatic 4175 cell line demonstrates extensive phosphorylation of p27 at the T157 and T198 sites. These sites have previously been implicated in the modulation of p27 localization and in the binding of p27 to RhoA. Thus, in the 4175 line, we demonstrate increased cytoplasmic p27, increased p27:RhoA binding, and impaired RhoA-ROCK signaling. This attenuation of RhoA activity correlates with increased migration (as measured by scratch assay) and increased invasion (as measured by modified transwell assay). Importantly, this hypermotility was abrogated by lentiviral knockdown of p27. Knockdown of p27 did not alter in vitro cell cycle profiles of either cell line as assessed by flow cytometry. The enhanced pulmonary tropism of the 4175 cell line was also abrogated in vivo following p27 knockdown: shRNAp27 reverts the lung metastatic phenotype of these cells back toward parental levels following tail vein injection in Balb/c nude female mice as assessed by IVIS imaging. Interestingly, orthotopic injection of these same lines, treated with the same lentivirus, into the mammary fat pad of Balb/c nude female mice produced a dramatically different result: shRNAp27 resulted in a significant increase in parental MDA-MB-231 tumor growth, while the metastatic derivative 4175 cells did not demonstrate a statistically meaningful change in orthotopic growth. These results indicate that, within the same cellular background, and in the same murine species, loss of p27 results in substantially different phenotypes depending upon the microenvironmental context: after tail vein injection cytoplasmic p27 plays a crucial role in promoting extravasation and colonization in the lung tissue; loss of p27 in this context impairs the metastatic process. Conversely, in the orthotopic environment, p27's pro-invasive role is not necessary to establish productive microenvironmental interactions and loss of p27 does not attenuate tumor growth. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2333. doi:10.1158/1538-7445.AM2011-2333</jats:p

Magnetic Polyacrylic Anion Exchange Resin: Preparation, Characterization and Adsorption Behavior of Humic Acid

Magnetic polyacrylic resin (R0) was prepared by suspension polymerization in the presence of γ-Fe₂O₃ and γ-methacryloxypropyltrimethoxysilane (γ-MPS). The obtained magnetic resin (R0) was modified sequentially by ammonolysis and alkylation to produce a magnetic weakly basic anion exchange resin (R1) and a magnetic strongly basic anion exchange resin (R2). Infrared (IR) spectra, elemental analysis, scanning electron microscopy (SEM), magnetization, Brunauer–Emmett–Teller (BET) surface area and chemical analysis were all determined to characterize these resins. The investigation of humic acid (HA) adsorption on R0, R1, and R2 showed that chemical interactions between the functional groups of the resin and HA were responsible for HA adsorption. The adsorption capacity of HA on R1 was ∼20%–60% less than that of R2, but the R1 resin was easier to regenerate with NaOH aqueous solution. A mixture of 1% NaCl and 1% NaOH was found to be highly efficient for the regeneration of both R1 and R2 resins, with desorption efficiencies measuring >90%. Pseudo-second-order model and Freundlich equation fit better for HA adsorption onto R1 and R2 than other typical kinetic and thermodynamic models, respectively. The effects of pH, coexisting salts (i.e., NaCl and Na₂SO₄), and recyclability were also assessed