ASU Scholarship Showcase

Neural progenitor cells (NPCs) derived from human pluripotent stem cells (hPSCs) are a multipotent cell population that is capable of nearly indefinite expansion and subsequent differentiation into the various neuronal and supporting cell types that comprise the CNS. However, current protocols for differentiating NPCs toward neuronal lineages result in a mixture of neurons from various regions of the CNS. In this study, we determined that endogenous WNT signaling is a primary contributor to the heterogeneity observed in NPC cultures and neuronal differentiation. Furthermore, exogenous manipulation of WNT signaling during neural differentiation, through either activation or inhibition, reduces this heterogeneity in NPC cultures, thereby promoting the formation of regionally homogeneous NPC and neuronal cultures. The ability to manipulate WNT signaling to generate regionally specific NPCs and neurons will be useful for studying human neural development and will greatly enhance the translational potential of hPSCs for neural-related therapies.

The threat of West Nile virus (WNV) epidemics with increasingly severe neuroinvasive infections demands the development and licensing of effective vaccines. To date, vaccine candidates based on inactivated, live-attenuated, or chimeric virus, and viral DNA and WNV protein subunits have been developed. Some have been approved for veterinary use or are under clinical investigation, yet no vaccine has been licensed for human use. Reaching the milestone of a commercialized human vaccine, however, may largely depend on the economics of vaccine production. Analysis suggests that currently only novel low-cost production technologies would allow vaccination to outcompete the cost of surveillance and clinical treatment. Here, we review progress using plants to address the economic challenges of WNV vaccine production. The advantages of plants as hosts for vaccine production in cost, speed and scalability, especially those of viral vector-based transient expression systems, are discussed. The progress in developing WNV subunit vaccines in plants is reviewed within the context of their expression, characterization, downstream processing, and immunogenicity in animal models. The development of vaccines based on enveloped and non-enveloped virus-like particles is also discussed. These advancements suggest that plants may provide a production platform that offers potent, safe and affordable human vaccines against WNV.

The termites evolved eusociality and complex societies before the ants, but have been studied much less. The recent publication of the first two termite genomes provides a unique comparative opportunity, particularly because the sequenced termites represent opposite ends of the social complexity spectrum. Zootermopsis nevadensis has simple colonies with totipotent workers that can develop into all castes (dispersing reproductives, nest-inheriting replacement reproductives, and soldiers). In contrast, the fungus-growing termite Macrotermes natalensis belongs to the higher termites and has very large and complex societies with morphologically distinct castes that are life-time sterile. Here we compare key characteristics of genomic architecture, focusing on genes involved in communication, immune defenses, mating biology and symbiosis that were likely important in termite social evolution. We discuss these in relation to what is known about these genes in the ants and outline hypothesis for further testing.

Although the majority of late-onset Alzheimer's disease (AD) patients are labeled sporadic, multiple genetic risk variants have been identified, the most powerful and prevalent of which is the e4 variant of the Apolipoprotein E (APOE) gene. Here, we generated human induced pluripotent stem cell (hiPSC) lines from the peripheral blood mononuclear cells (PBMCs) of a clinically diagnosed AD patient [ASUi003-A] and a non-demented control (NDC) patient [ASUi004-A] homozygous for the APOE4 risk allele. These hiPSCs maintained their original genotype, expressed pluripotency markers, exhibited a normal karyotype, and retained the ability to differentiate into cells representative of the three germ layers.

Nonsense-mediated RNA decay (NMD) is a highly conserved pathway that selectively degrades specific subsets of RNA transcripts. Here, we provide evidence that NMD regulates early human developmental cell fate. We found that NMD factors tend to be expressed at higher levels in human pluripotent cells than in differentiated cells, raising the possibility that NMD must be downregulated to permit differentiation. Loss- and gain-of-function experiments in human embryonic stem cells (hESCs) demonstrated that, indeed, NMD downregulation is essential for efficient generation of definitive endoderm. RNA-seq analysis identified NMD target transcripts induced when NMD is suppressed in hESCs, including many encoding signaling components. This led us to test the role of TGF-β and BMP signaling, which we found NMD acts through to influence definitive endoderm versus mesoderm fate. Our results suggest that selective RNA decay is critical for specifying the developmental fate of specific human embryonic cell lineages.

Previously, our group engineered a plant-derived monoclonal antibody (MAb) (pHu-E16) that efficiently treated West Nile virus (WNV) infection in mice. In this study, we developed several pHu-E16 variants to improve its efficacy. These variants included a single-chain variable fragment (scFv) of pHu-E16 fused to the heavy chain (HC) constant domains (CH1-3) of human IgG (pHu-E16scFv-CH1-3) and a tetravalent molecule (Tetra pHu-E16) assembled from pHu-E16scFv-CH1-3 with a second pHu-E16scFv fused to the light chain (LC) constant region. pHu-E16scFv-CH1-3 and Tetra pHu-E16 were efficiently expressed and assembled in plants. To assess the impact of differences in N-linked glycosylation on pHu-E16 variant assembly and function, we expressed additional pHu-E16 variants with various combinations of HC and LC components.

Our study revealed that proper pairing of HC and LC was essential for the complete N-glycan processing of antibodies in both plant and animal cells. Associated with their distinct N-glycoforms, pHu-E16, pHu-E16scFv-CH1-3 and Tetra pHu-E16 exhibited differential binding to C1q and specific Fcγ receptors (FcγR). Notably, none of the plant-derived Hu-E16 variants showed antibody-dependent enhancement (ADE) activity in CD32A+ human cells, suggesting the potential of plant-produced antibodies to minimize the adverse effect of ADE. Importantly, all plant-derived MAb variants exhibited at least equivalent in vitro neutralization and in vivo protection in mice compared to mammalian cell-produced Hu-E16. This study demonstrates the capacity of plants to express and assemble a large, complex and functional IgG-like tetravalent mAb variant and also provides insight into the relationship between MAb N-glycosylation, FcγR and C1q binding, and ADE. These new insights may allow the development of safer and cost effective MAb-based therapeutics for flaviviruses, and possibly other pathogens.

Background: Mutual policing is an important mechanism for reducing conflict in cooperative groups. In societies of ants, bees, and wasps, mutual policing of worker reproduction can evolve when workers are more closely related to the queen's sons than to the sons of workers or when the costs of worker reproduction lower the inclusive fitness of workers. During colony growth, relatedness within the colony remains the same, but the costs of worker reproduction may change. The costs of worker reproduction are predicted to be greatest in incipient colonies. If the costs associated with worker reproduction outweigh the individual direct benefits to workers, policing mechanisms as found in larger colonies may be absent in incipient colonies.

Results: We investigated policing behavior across colony growth in the ant 'Camponotus floridanus.' In large colonies of this species, worker reproduction is policed by the destruction of worker-laid eggs. We found workers from incipient colonies do not exhibit policing behavior, and instead tolerate all conspecific eggs. The change in policing behavior is consistent with changes in egg surface hydrocarbons, which provide the informational basis for policing; eggs laid by queens from incipient colonies lack the characteristic hydrocarbons on the surface of eggs laid by queens from large colonies, making them chemically indistinguishable from worker-laid eggs. We also tested the response to fertility information in the context of queen tolerance. Workers from incipient colonies attacked foreign queens from large colonies; whereas workers from large colonies tolerated such queens. Workers from both incipient and large colonies attacked foreign queens from incipient colonies.

Conclusions: Our results provide novel insights into the regulation of worker reproduction in social insects at both the proximate and ultimate levels. At the proximate level, our results show that mechanisms of social regulation, such as the response to fertility signals, change dramatically over a colony's life cycle. At the ultimate level, our results emphasize the importance of factors besides relatedness in predicting the level of conflict within a colony. Our results also suggest policing may not be an important regulatory force at every stage of colony development. Changes relating to the life cycle of the colony are sufficient to account for major differences in social regulation in an insect colony. Mechanisms of conflict mediation observed in one phase of a social group's development cannot be generalized to all stages.

In the decade since Yamanaka and colleagues described methods to reprogram somatic cells into a pluripotent state, human induced pluripotent stem cells (hiPSCs) have demonstrated tremendous promise in numerous disease modeling, drug discovery, and regenerative medicine applications. More recently, the development and refinement of advanced gene transduction and editing technologies have further accelerated the potential of hiPSCs. In this review, we discuss the various gene editing technologies that are being implemented with hiPSCs. Specifically, we describe the emergence of technologies including zinc-finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 that can be used to edit the genome at precise locations, and discuss the strengths and weaknesses of each of these technologies. In addition, we present the current applications of these technologies in elucidating the mechanisms of human development and disease, developing novel and effective therapeutic molecules, and engineering cell-based therapies. Finally, we discuss the emerging technological advances in targeted gene editing methods.

Adult and pluripotent stem cells represent a ready supply of cellular raw materials that can be used to generate the functionally mature cells needed to replace damaged or diseased heart tissue. However, the use of stem cells for cardiac regenerative therapies is limited by the low efficiency by which stem cells are differentiated in vitro to cardiac lineages as well as the inability to effectively deliver stem cells and their derivatives to regions of damaged myocardium. In this review, we discuss the various biomaterial-based approaches that are being implemented to direct stem cell fate both in vitro and in vivo. First, we discuss the stem cell types available for cardiac repair and the engineering of naturally and synthetically derived biomaterials to direct their in vitro differentiation to the cell types that comprise heart tissue. Next, we describe biomaterial-based approaches that are being implemented to enhance the in vivo integration and differentiation of stem cells delivered to areas of cardiac damage. Finally, we present emerging trends of using stem cell-based biomaterial approaches to deliver pro-survival factors and fully vascularized tissue to the damaged and diseased cardiac tissue.

The field of tissue engineering entered a new era with the development of human pluripotent stem cells (hPSCs), which are capable of unlimited expansion whilst retaining the potential to differentiate into all mature cell populations. However, these cells harbor significant risks, including tumor formation upon transplantation. One way to mitigate this risk is to develop expandable progenitor cell populations with restricted differentiation potential. Here, we used a cellular microarray technology to identify a defined and optimized culture condition that supports the derivation and propagation of a cell population with mesodermal properties. This cell population, referred to as intermediate mesodermal progenitor (IMP) cells, is capable of unlimited expansion, lacks tumor formation potential, and, upon appropriate stimulation, readily acquires properties of a sub-population of kidney cells. Interestingly, IMP cells fail to differentiate into other mesodermally-derived tissues, including blood and heart, suggesting that these cells are restricted to an intermediate mesodermal fate.