COPD Subtypes Are Differentially Associated with Cardiovascular Events and COPD Exacerbations.

BACKGROUND: The coronary artery calcium score (CACS) and ratio of the pulmonary artery to aorta diameters (PA:A ratio) measured from chest CT scans have been established as predictors of cardiovascular events and chronic obstructive pulmonary disease (COPD) exacerbations, respectively. However, little is known about the reciprocal relationship between these predictors and outcomes. Furthermore, the prognostic implications of COPD subtypes on clinical outcomes remain insufficiently characterized. RESEARCH QUESTION: How can these two chest CT-derived parameters predict subsequent cardiovascular events and COPD exacerbations in different COPD subtypes? STUDY DESIGN AND METHODS: Using COPDGene study data, we assessed prospective cardiovascular disease (CVD) and COPD exacerbation risk in COPD subjects (Global Initiative for Chronic Obstructive Lung Disease spirometric grades 2-4), focusing on CACS and PA:A ratio at study enrollment, with logistic regression models. These outcomes were analyzed in three COPD subtypes: 1,042 Non-emphysema-predominant COPD (NEPD; low attenuation area at -950 Hounsfield units [LAA-950]<5%), 1,324 Emphysema-predominant COPD (EPD; LAA-950≥10%), and 465 Intermediate Emphysema COPD (IE; 5≤LAA-950<10%). RESULTS: Our study indicated significantly higher overall risk for cardiovascular events in subjects with higher CACS (≥median; Odds Ratio (OR): 1.61, 95% Confidence Interval (CI)=1.30-2.00) and increased COPD exacerbations in those with higher PA:A ratios (≥1; OR: 1.80, 95% CI=1.46-2.23). Notably, NEPD subjects showed a stronger association between these indicators and clinical events compared to EPD (with CACS/CVD, NEPD vs. EPD, OR 2.02 vs. 1.41; with PA:A ratio/COPD exacerbation, NEPD vs. EPD, OR 2.50 vs. 1.65); the difference in odds ratios between COPD subtypes was statistically significant for CACS/CVD. INTERPRETATION: Two chest CT parameters, CACS and PA:A ratio, hold distinct predictive values for cardiovascular events and COPD exacerbations that are influenced by specific COPD subtypes. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT00608764.

Promoting Human Intestinal Organoid Formation and Stimulation Using Piezoelectric Nanofiber Matrices.

Human organoid model systems have changed the landscape of developmental biology and basic science. They serve as a great tool for human specific interrogation. In order to advance our organoid technology, we aimed to test the compatibility of a piezoelectric material with organoid generation, because it will create a new platform with the potential for sensing and actuating organoids in physiologically relevant ways. We differentiated human pluripotent stem cells into spheroids following the traditional human intestinal organoid (HIO) protocol atop a piezoelectric nanofiber scaffold. We observed that exposure to the biocompatible piezoelectric nanofibers promoted spheroid morphology three days sooner than with the conventional methodology. At day 28 of culture, HIOs grown on the scaffold appeared similar. Both groups were readily transplantable and developed well-organized laminated structures. Graft sizes between groups were similar. Upon characterizing the tissue further, we found no detrimental effects of the piezoelectric nanofibers on intestinal patterning or maturation. Furthermore, to test the practical feasibility of the material, HIOs were also matured on the nanofiber scaffolds and treated with ultrasound, which lead to increased cellular proliferation which is critical for organoid development and tissue maintenance. This study establishes a proof of concept for integrating piezoelectric materials as a customizable platform for on-demand electrical stimulation of cells using remote ultrasonic waveforms in regenerative medicine.

RFX6 regulates human intestinal patterning and function upstream of PDX1.

The gastrointestinal (GI) tract is complex and consists of multiple organs with unique functions. Rare gene variants can cause congenital malformations of the human GI tract, although the molecular basis of these has been poorly studied. We identified a patient with compound-heterozygous variants in RFX6 presenting with duodenal malrotation and atresia, implicating RFX6 in development of the proximal intestine. To identify how mutations in RFX6 impact intestinal patterning and function, we derived induced pluripotent stem cells from this patient to generate human intestinal organoids (HIOs). We identified that the duodenal HIOs and human tissues had mixed regional identity, with gastric and ileal features. CRISPR-mediated correction of RFX6 restored duodenal identity. We then used gain- and loss-of-function and transcriptomic approaches in HIOs and Xenopus embryos to identify that PDX1 is a downstream transcriptional target of RFX6 required for duodenal development. However, RFX6 had additional PDX1-independent transcriptional targets involving multiple components of signaling pathways that are required for establishing early regional identity in the GI tract. In summary, we have identified RFX6 as a key regulator in intestinal patterning that acts by regulating transcriptional and signaling pathways.

Deciphering Endothelial and Mesenchymal Organ Specification in Vascularized Lung and Intestinal Organoids.

To investigate the co-development of vasculature, mesenchyme, and epithelium crucial for organogenesis and the acquisition of organ-specific characteristics, we constructed a human pluripotent stem cell-derived organoid system comprising lung or intestinal epithelium surrounded by organotypic mesenchyme and vasculature. We demonstrated the pivotal role of co-differentiating mesoderm and endoderm via precise BMP regulation in generating multilineage organoids and gut tube patterning. Single-cell RNA-seq analysis revealed organ specificity in endothelium and mesenchyme, and uncovered key ligands driving endothelial specification in the lung (e.g., WNT2B and Semaphorins) or intestine (e.g., GDF15). Upon transplantation under the kidney capsule in mice, these organoids further matured and developed perfusable human-specific sub-epithelial capillaries. Additionally, our model recapitulated the abnormal endothelial-epithelial crosstalk in patients with FOXF1 deletion or mutations. Multilineage organoids provide a unique platform to study developmental cues guiding endothelial and mesenchymal cell fate determination, and investigate intricate cell-cell communications in human organogenesis and disease. Highlights: BMP signaling fine-tunes the co-differentiation of mesoderm and endoderm.The cellular composition in multilineage organoids resembles that of human fetal organs.Mesenchyme and endothelium co-developed within the organoids adopt organ-specific characteristics.Multilineage organoids recapitulate abnormal endothelial-epithelial crosstalk in FOXF1-associated disorders.

Eicosatetraynoic Acid Regulates Pro-Fibrotic Pathways in an Induced Pluripotent Stem Cell Derived Macrophage:Human Intestinal Organoid Model of Crohn's Disease.

Background and Aims: We previously identified small molecules predicted to reverse an ileal gene signature for future Crohn's Disease (CD) strictures. Here we used a new human intestinal organoid (HIO) model system containing macrophages to test a lead candidate, eicosatetraynoic acid (ETYA). Methods: Induced pluripotent stem cell lines (iPSC) were derived from CD patients and differentiated into macrophages and HIOs. Macrophages and macrophage:HIO co-cultures were exposed to lipopolysaccharide (LPS) with and without ETYA pre-treatment. Cytospin and flow cytometry characterized macrophage morphology and activation markers, and RNA sequencing defined the global pattern of macrophage gene expression. TaqMan Low Density Array, Luminex multiplex assay, immunohistologic staining, and sirius red polarized light microscopy were performed to measure macrophage cytokine production and HIO pro-fibrotic gene expression and collagen content. Results: iPSC-derived macrophages exhibited morphology similar to primary macrophages and expressed inflammatory macrophage cell surface markers including CD64 and CD68. LPS-stimulated macrophages expressed a global pattern of gene expression enriched in CD ileal inflammatory macrophages and matrisome secreted products, and produced cytokines and chemokines including CCL2, IL1B, and OSM implicated in refractory disease. ETYA suppressed CD64 abundance and pro-fibrotic gene expression pathways in LPS stimulated macrophages. Co-culture of LPS-primed macrophages with HIO led to up-regulation of fibroblast activation genes including ACTA2 and COL1A1 , and an increase in HIO collagen content. ETYA pre-treatment prevented pro-fibrotic effects of LPS-primed macrophages. Conclusions: ETYA inhibits pro-fibrotic effects of LPS-primed macrophages upon co-cultured HIO. This model may be used in future untargeted screens for small molecules to treat refractory CD.