Gude, N. | bioRxiv Science

bioRxiv2021

Fundamentals of Vaping-Associated Pulmonary Injury Leading to Severe Respiratory Distress

Vaping of flavored liquids has been touted as safe alternative to traditional cigarette smoking with decreased health risks. The popularity of vaping has dramatically increased over the last decade, particularly among teenagers who incorporate vaping into their daily life as a social activity. Despite widespread and increasing adoption of vaping among young adults there is little information on long term consequences of vaping and potential health risks. This study demonstrates Vaping-Induced Pulmonary Injury (VAPI) using commercial JUUL pens with flavored vape juice using an inhalation exposure murine model. Profound pathological changes to upper airway, lung tissue architecture, and cellular structure are evident within 9 weeks of exposure. Marked histologic changes include increased parenchyma tissue density, cellular infiltrates proximal to airway passages, alveolar rarefaction, increased collagen deposition, and bronchial thickening with elastin fiber disruption. Transcriptional reprogramming includes significant changes to gene families coding for xenobiotic response, glycerolipid metabolic processes, and oxidative stress. Cardiac contractile performance for systemic output is moderately but significantly impaired, and the shows severe pulmonary side structural remodeling with chamber enlargement. This VAPI model with pulmonary circuit failure demonstrates mechanistic underpinnings of vaping-related pathologic injury.

molecular biology↗

bioRxiv2019

Enhancing Myocardial Repair with CardioClusters

BackgroundCellular therapy to treat heart failure is an ongoing focus of intense research and development, but progress has been frustratingly slow due to limitations of current approaches. Engineered augmentation of established cellular effectors overcomes impediments, enhancing reparative activity with improved outcomes relative to conventional techniques. Such next generation implementation includes delivery of combinatorial cell populations exerting synergistic effects. Concurrent isolation and expansion of three distinct cardiac-derived interstitial cell types from human heart tissue, as previously reported by our group, prompted design of a three-dimensional (3D) structure that maximizes cellular interaction, allows for defined cell ratios, controls size, enables injectability, and minimizes cell losses upon delivery. MethodsThree distinct populations of human cardiac interstitial cells including mesenchymal stem cells (MSCs), endothelial progenitor cells (EPCs), and c-Kit+ cardiac interstitial cells (cCICs) when cultured together spontaneously form scaffold-free 3D microenvironments termed CardioClusters. Biological consequences of CardioCluster formation were assessed by multiple assays including single cells RNA-Seq transcriptional profiling. Protective effects of CardioClusters in vitro were measured using cell culture models for oxidative stress and myocardial ischemia in combination with freshly isolated neonatal rat ventricular myocytes. Long-term impact of adoptively transferred CardioClusters upon myocardial structure and function in a xenogenic model of acute infarction using NODscid mice was assessed over a longitudinal time course of 20-weeks. ResultsCardioCluster design enables control over composite cell types, cell ratios, size, and preservation of structural integrity during delivery. Profound changes for biological properties of CardioClusters relative to constituent parental cell populations include enhanced expression of stem cell-relevant factors, adhesion/extracellular-matrix molecules, and cytokines. The CardioCluster 3D microenvironment maximizes cellular interaction while maintaining a more native transcriptome similar to endogenous cardiac cells. CardioCluster delivery improves cell retention following intramyocardial injection with preservation of long-term cardiac function relative to monolayer-cultured cells when tested in an experimental murine infarction model followed for up to 20 weeks post-challenge. CardioCluster-treated hearts show increases in capillary density, preservation of cardiomyocyte size, and reduced scar size indicative of blunting pathologic infarction injury. ConclusionsCardioClusters are a novel next generation development and delivery approach for cellular therapeutics that potentiate beneficial activity and enhance protective effects of human cardiac interstitial cell mixed populations. CardioClusters utilization in this preclinical setting establishes fundamental methodologic and biologic insights, laying the framework for optimization of CardioCluster design to provide greater efficacy in cell-based therapeutic interventions intended to mitigate cardiomyopathic damage.

cell biology↗

bioRxiv2019

Tetraploidy in rodent cardiac stem cells confers enhanced biological properties

Ploidy for cardiomyocytes is well described but remains obscure in cardiac interstitial cells (CICs). Ploidy of c-kit+CICs were assessed using a combination of confocal, karyotypic, and flow cytometric assessments coupled with molecular and bioinformatic analyses. Fundamental differences were found between cultured rodent (rat, mouse) c-kit+CICs possessing mononuclear tetraploid (4n) content versus large mammal (human, swine) with mononuclear diploid (2n) content. In-situ analysis, confirmed with fresh isolates, revealed diploid content in c-kit+CICs from human and a mixture of diploid and tetraploid nuclei in mouse. Molecular assessment of the p53 signaling pathway provides a plausible explanation for escape from replicative senescence in rodent but not human ckit+CICs. Single cell transcriptional profiling reveals distinctions between diploid versus tetraploid populations in mouse ckit+CICs, alluding to functional divergences. Collectively, these data reveal fundamental species-specific biological differences in c-kit+CICs that could account for challenges in extrapolation of myocardial preclinical studies from rodent to large animal models.

cell biology↗