Model For Study Of Pathophysiology Research Articles

A 3-D human model of complex cardiac arrhythmias

Cardiac arrhythmias impact over 12 million people globally, with an increasing incidence of acquired arrhythmias. Although animal models have shed light onto fundamental arrhythmic mechanisms, species-specific differences and ethical concerns remain. Current human models using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) either lack the higher order tissue organization of the heart or implement unreliable arrhythmia induction techniques. Our goal was to develop a robust model of acquired arrhythmia by disrupting cardiomyocyte cell-cell signaling – one of the hallmarks of complex arrhythmias. Human 3D microtissues were generated by seeding hydrogel-embedded hiPSC-CMs and cardiac fibroblasts into an established microwell system designed to enable active and passive force assessment. Cell-cell signaling was disrupted using methyl-beta cyclodextrin (MBCD), previously shown to disassemble cardiac gap junctions. We demonstrate that arrhythmias were progressive and present in all microtissues within 5 days of treatment. Arrhythmic tissues exhibited reduced conduction velocity, an increased number of distinct action potentials, and reduced action potential cycle length. Arrhythmic tissues also showed significant reduction in contractile force generation, increased beating frequency, and increased passive tension and collagen deposition, in line with fibrosis. A subset of tissues with more complex arrhythmias exhibited 3D spatial differences in action potential propagation. Pharmacological and electrical defibrillation was successful. Transcriptomic data indicated an enrichment of genes consistent with cardiac arrhythmias. MBCD removal reversed the arrhythmic phenotype, resulting in synchronicity despite not resolving fibrosis. This innovative & reliable human-relevant 3D acquired arrhythmia model shows potential for improving our understanding of arrhythmic action potential conduction and furthering therapeutic development. Statement of significanceThis work describes a 3D human model of cardiac arrhythmia-on-a-chip with high reproducibility, fidelity, and extensive functional applicability. To mimic in vivo conditions, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and cardiac fibroblasts from healthy controls were combined in a biocompatible fibrin hydrogel and seeded between two deflectable polymeric rods. Using the innate functional properties of this 3D model as well as advanced optical imaging techniques we demonstrated dramatic changes in contraction rate, synchronicity, and electrophysiological conduction in arrhythmic tissues relative to controls. Taken together, these data demonstrate the distinctive potential of this new model for pathophysiological studies, and for arrhythmia drug testing applications.

A Nonhuman Primate Model of Stable Urinary Sphincter Deficiency

The pathophysiology of urinary sphincter deficiency in women remains incompletely understood and current treatment options have limitations. Female nonhuman primates may represent a relevant animal model for studies of pathophysiology and treatment interventions because of their human-like reproductive and age associated stages of life (premenopause, perimenopause and postmenopause), lower urinary tract structure and bipedal posture. We developed and characterized a nonhuman primate model of defined injury to the urethral sphincter complex. We used 22 adult female cynomolgus monkeys in which injury to the sphincter complex was created by cauterizing and then transecting its pudendal innervation. Urodynamic studies were performed before and during pudendal and hypogastric nerve stimulation at baseline, and 3, 6 and 12 months after injury. We also analyzed sphincter structure in vivo by cystourethrography, and ex vivo by quantitative histology and immunohistochemistry at these time points. Injury produced a 47% to 50% decrease in maximal urethral pressure (vs baseline p <0.05). It also abolished the increase in maximal urethral pressure in response to pudendal and hypogastric nerve stimulation (vs baseline p >0.05), which persisted more than 12 months after injury. Urodynamic changes were consistent with decreased skeletal and smooth muscle content, decreased nerve responses and an associated decrease in somatic and adrenergic innervation in the sphincter complex. These structural and urodynamic changes are consistent with those in patients with stress urinary incontinence. They support the usefulness of nonhuman primates as translatable surrogates for pathophysiological studies of urinary sphincter deficiency and testing novel therapies for that condition.

Mouse model of Sanfilippo syndrome type B produced by targeted disruption of the gene encoding alpha-N-acetylglucosaminidase.

The Sanfilippo syndrome type B is an autosomal recessive disorder caused by mutation in the gene (NAGLU) encoding alpha-N-acetylglucosaminidase, a lysosomal enzyme required for the stepwise degradation of heparan sulfate. The most serious manifestations are profound mental retardation, intractable behavior problems, and death in the second decade. To generate a model for studies of pathophysiology and of potential therapy, we disrupted exon 6 of Naglu, the homologous mouse gene. Naglu-/- mice were healthy and fertile while young and could survive for 8-12 mo. They were totally deficient in alpha-N-acetylglucosaminidase and had massive accumulation of heparan sulfate in liver and kidney as well as secondary changes in activity of several other lysosomal enzymes in liver and brain and elevation of gangliosides G(M2) and G(M3) in brain. Vacuolation was seen in many cells, including macrophages, epithelial cells, and neurons, and became more prominent with age. Although most vacuoles contained finely granular material characteristic of glycosaminoglycan accumulation, large pleiomorphic inclusions were seen in some neurons and pericytes in the brain. Abnormal hypoactive behavior was manifested by 4.5-mo-old Naglu-/- mice in an open field test; the hyperactivity that is characteristic of affected children was not observed even in younger mice. In a Pavlovian fear conditioning test, the 4.5-mo-old mutant mice showed normal response to context, indicating intact hippocampal-dependent learning, but reduced response to a conditioning tone, perhaps attributable to hearing impairment. The phenotype of the alpha-N-acetylglucosaminidase-deficient mice is sufficiently similar to that of patients with the Sanfilippo syndrome type B to make these mice a good model for study of pathophysiology and for development of therapy.