Multi-lineage heart-chip models drug cardiotoxicity and enhances maturation of human stem cell-derived cardiovascular cells.

Cardiovascular toxicity causes adverse drug reactions and may lead to drug removal from the pharmaceutical market. Cancer therapies can induce life-threatening cardiovascular side effects such as arrhythmias, muscle cell death, or vascular dysfunction. New technologies have enabled cardiotoxic compounds to be identified earlier in drug development. Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs) and vascular endothelial cells (ECs) can screen for drug-induced alterations in cardiovascular cell function and survival. However, most existing hiPSC models for cardiovascular drug toxicity utilize two-dimensional, immature cells grown in static culture. Improved in vitro models to mechanistically interrogate cardiotoxicity would utilize more adult-like, mature hiPSC-derived cells in an integrated system whereby toxic drugs and protective agents can flow between hiPSC-ECs that represent systemic vasculature and hiPSC-CMs that represent heart muscle (myocardium). Such models would be useful for testing the multi-lineage cardiotoxicities of chemotherapeutic drugs such as VEGFR2/PDGFR-inhibiting tyrosine kinase inhibitors (VPTKIs). Here, we develop a multi-lineage, fully-integrated, cardiovascular organ-chip that can enhance hiPSC-EC and hiPSC-CM functional and genetic maturity, model endothelial barrier permeability, and demonstrate long-term functional stability. This microfluidic organ-chip harbors hiPSC-CMs and hiPSC-ECs on separate channels that can be subjected to active fluid flow and rhythmic biomechanical stretch. We demonstrate the utility of this cardiovascular organ-chip as a predictive platform for evaluating multi-lineage VPTKI toxicity. This study may lead to the development of new modalities for the evaluation and prevention of cancer therapy-induced cardiotoxicity.

Human iPSC-derived neural progenitor cells secreting GDNF provide protection in rodent models of ALS and retinal degeneration.

Human induced pluripotent stem cells (iPSCs) are a renewable cell source that can be differentiated into neural progenitor cells (iNPCs) and transduced with glial cell line-derived neurotrophic factor (iNPC-GDNFs). The goal of the current study is to characterize iNPC-GDNFs and test their therapeutic potential and safety. Single-nuclei RNA-seq show iNPC-GDNFs express NPC markers. iNPC-GDNFs delivered into the subretinal space of the Royal College of Surgeons rodent model of retinal degeneration preserve photoreceptors and visual function. Additionally, iNPC-GDNF transplants in the spinal cord of SOD1G93A amyotrophic lateral sclerosis (ALS) rats preserve motor neurons. Finally, iNPC-GDNF transplants in the spinal cord of athymic nude rats survive and produce GDNF for 9 months, with no signs of tumor formation or continual cell proliferation. iNPC-GDNFs survive long-term, are safe, and provide neuroprotection in models of both retinal degeneration and ALS, indicating their potential as a combined cell and gene therapy for various neurodegenerative diseases.

Activated Bone Marrow-Derived Macrophages Eradicate Alzheimer's-Related Aß42 Oligomers and Protect Synapses.

Impaired synaptic integrity and function due to accumulation of amyloid ß-protein (Aß42) oligomers is thought to be a major contributor to cognitive decline in Alzheimer's disease (AD). However, the exact role of Aß42 oligomers in synaptotoxicity and the ability of peripheral innate immune cells to rescue synapses remain poorly understood due to the metastable nature of oligomers. Here, we utilized photo-induced cross-linking to stabilize pure oligomers and study their effects vs. fibrils on synapses and protection by Aß-phagocytic macrophages. We found that cortical neurons were more susceptible to Aß42 oligomers than fibrils, triggering additional neuritic arborization retraction, functional alterations (hyperactivity and spike waveform), and loss of VGluT1- and PSD95-excitatory synapses. Co-culturing neurons with bone marrow-derived macrophages protected synapses against Aß42 fibrils; moreover, immune activation with glatiramer acetate (GA) conferred further protection against oligomers. Mechanisms involved increased Aß42 removal by macrophages, amplified by GA stimulation: fibrils were largely cleared through intracellular CD36/EEA1+-early endosomal proteolysis, while oligomers were primarily removed via extracellular/MMP-9 enzymatic degradation. In vivo studies in GA-immunized or CD115+-monocyte-grafted APPSWE/PS1ΔE9-transgenic mice followed by pre- and postsynaptic analyses of entorhinal cortex and hippocampal substructures corroborated our in vitro findings of macrophage-mediated synaptic preservation. Together, our data demonstrate that activated macrophages effectively clear Aß42 oligomers and rescue VGluT1/PSD95 synapses, providing rationale for harnessing macrophages to treat AD.

Removal of endogenous neuromodulators in a small motor network enhances responsiveness to neuromodulation.

We studied the changes in sensitivity to a peptide modulator, crustacean cardioactive peptide (CCAP), as a response to loss of endogenous modulation in the stomatogastric ganglion (STG) of the crab Cancer borealis Our data demonstrate that removal of endogenous modulation for 24 h increases the response of the lateral pyloric (LP) neuron of the STG to exogenously applied CCAP. Increased responsiveness is accompanied by increases in CCAP receptor (CCAPr) mRNA levels in LP neurons, requires de novo protein synthesis, and can be prevented by coincubation for the 24-h period with exogenous CCAP. These results suggest that there is a direct feedback from loss of CCAP signaling to the production of CCAPr that increases subsequent response to the ligand. However, we also demonstrate that the modulator-evoked membrane current (IMI) activated by CCAP is greater in magnitude after combined loss of endogenous modulation and activity compared with removal of just hormonal modulation. These results suggest that both receptor expression and an increase in the target conductance of the CCAP G protein-coupled receptor are involved in the increased response to exogenous hormone exposure following experimental loss of modulation in the STG.NEW & NOTEWORTHY The nervous system shows a tremendous amount of plasticity. More recently there has been an appreciation for compensatory actions that stabilize output in the face of perturbations to normal activity. In this study we demonstrate that neurons of the crustacean stomatogastric ganglion generate apparent compensatory responses to loss of peptide neuromodulation, adding to the repertoire of mechanisms by which the stomatogastric nervous system can regulate and stabilize its own output.

Neuropeptide receptor transcript expression levels and magnitude of ionic current responses show cell type-specific differences in a small motor circuit.

We studied the relationship between neuropeptide receptor transcript expression and current responses in the stomatogastric ganglion (STG) of the crab, Cancer borealis. We identified a transcript with high sequence similarity to crustacean cardioactive peptide (CCAP) receptors in insects and mammalian neuropeptide S receptors. This transcript was expressed throughout the nervous system, consistent with the role of CCAP in a range of different behaviors. In the STG, single-cell qPCR showed expression in only a subset of neurons. This subset had previously been shown to respond to CCAP with the activation of a modulator-activated inward current (IMI), with one exception. In the one cell type that showed expression but no IMI responses, we found CCAP modulation of synaptic currents. Expression levels within STG neuron types were fairly variable, but significantly different between some neuron types. We tested the magnitude and concentration dependence of IMI responses to CCAP application in two identified neurons, the lateral pyloric (LP) and the inferior cardiac (IC) neurons. LP had several-fold higher expression and showed larger current responses. It also was more sensitive to low CCAP concentrations and showed saturation at lower concentrations, as sigmoid fits showed smaller EC50 values and steeper slopes. In addition, occlusion experiments with proctolin, a different neuropeptide converging onto IMI, showed that saturating concentrations of CCAP activated all available IMI in LP, but only approximately two-thirds in IC, the neuron with lower receptor transcript expression. The implications of these findings for comodulation are discussed.