Biology subjects

Conchola, A. S.

Publications and source records attributed to Conchola, A. S..

7 recordsLinked to original sources

bioRxiv2026

Bidirectional fibrogenic cross-talk revealed in a human iPSC-derived epithelial-mesenchymal co-culture model of pulmonary fibrosis

Pulmonary fibrosis (PF) can arise from mutations in alveolar epithelial type 2 (AT2) cell-specific genes, but manifests in fibrotic activation of mesenchymal cells, thus involving fibrogenic epithelial-mesenchymal crosstalk. The ligand-receptor interactions underlying the onset and early progression of PF remain poorly understood. Induced pluripotent stem cell (iPSC)-derived models are powerful tools to study respiratory diseases, yet are currently limited to reductionist single lineage epithelial models or multi-lineage systems that lack purity and lung-specificity of the mesenchyme. Here we generate a human iPSC line carrying both a lung mesenchyme-specific reporter (TBX4-LERtdTomato) and a reporter for mesenchymal activation/differentiation (ACTA2GFP). Applying this line, we develop a directed differentiation protocol capable of generating cells that express key molecular and functional features of primary human developing lung mesenchyme across multiple iPSC genetic backgrounds. We then establish co-cultures of these iPSC-derived lung mesenchymal cells (iLM) with patient-specific iPSC-derived alveolar epithelial type 2 cells (iAT2s) carrying an SFTPCI73T mutation as a model for PF. We find increased expression of fibrotic markers in co-cultures with mutant iAT2s as compared to co-cultures with gene-corrected iAT2s. Moreover, mutant iAT2s express markers of alveolar-basal intermediate (ABI) cells only in the presence of iLM, suggesting that bidirectional crosstalk promotes this aberrant cell state. We identify ligand-receptor pairs enriched in co-cultures with mutant iAT2s, including TGF{beta}, multiple integrins, and additional genes that have not been previously linked to PF. Finally, we show that small molecule-mediated inhibition of TGF{beta} or integrins V{beta}1/V{beta}6 attenuates both fibrotic mesenchymal activation and the presence of ABI cells in iLM/iAT2 co-cultures. Thus, we have established a human iPSC-derived co-culture system that recapitulates key molecular hallmarks of bidirectional fibrogenic epithelial-mesenchymal crosstalk in pulmonary fibrosis, and enables the identification and study of potentially druggable pathways involved in disease initiation and progression.

molecular biology↗

bioRxiv2024

A spatially-resolved blueprint of the developing human lung reveals a basal cell population regulated by a WNT-driven niche

Organs are composed of diverse cell types that change across space and time during development. To interrogate this diversity, we micro-dissected developing human lungs along the proximal-distal axis during the late pseudoglandular stage and generated an integrated analysis of single-nucleus sequencing and spatial transcriptomics, creating a cellularly-resolved atlas of the lung. These rich datasets revealed positional niches and cellular heterogeneity along the proximal-distal axis, including the identification of a unique population of TP63+ basal cells, the primary stem cell of the airway, marked by expression of LGR5 and LGR6. Analysis of the LGR5+ basal cell niche and functional experiments with primary organoid models suggest a tonic level of WNT pathway activity in LGR5+ basal cells that is potentiated by mesenchyme-derived R-SPONDIN. We found that basal cell self-renewal is enhanced by WNT activity, suggesting that the WNT pathway plays a previously unappreciated but critical role in airway stem cell maintenance during human development. These results enhance our fundamental understanding of the positional and cellular heterogeneity in the developing human lungs and begin to reveal unique niches that maintain homeostasis throughout the lung.

developmental biology↗

bioRxiv2023

Coordinated differentiation of human intestinal organoids with functional enteric neurons and vasculature

Human intestinal organoids (HIOs) derived from human pluripotent stem cells co-differentiate both epithelial and mesenchymal lineages in vitro but lack important cell types such as neurons, endothelial cells, and smooth muscle. Here, we report an in vitro method to derive HIOs with epithelium, mesenchyme, enteric neuroglial populations, endothelial cells, and organized smooth muscle in a single differentiation, without the need for co-culture. When transplanted into a murine host, these populations expand and organize to support organoid maturation and function. Functional experiments demonstrate enteric nervous system function, with HIOs undergoing peristaltic-like contractions, suggesting the development of a functional neuromuscular unit. HIOs also form functional vasculature, demonstrated in vitro using microfluidic devices to introduce vascular-like flow, and in vivo following transplantation, where HIO endothelial cells anastomose with host vasculature. Collectively, we report an in vitro model of the human gut that simultaneously co-differentiates epithelial, stromal, endothelial, neural, and organized muscle populations.

bioengineering↗

bioRxiv2023

Opposing roles for TGFbeta and BMP-signaling during nascent alveolar differentiation in the developing human lung

Alveolar type 2 (AT2) cells function as stem cells in the adult lung and aid in repair after injury. The current study aimed to understand the signaling events that control differentiation of this therapeutically relevant cell type during human development. Using lung explant and organoid models, we identified opposing effects of TGF{beta}- and BMP-signaling, where inhibition of TGF{beta}- and activation of BMP-signaling in the context of high WNT- and FGF-signaling efficiently differentiated early lung progenitors into AT2-like cells in vitro. AT2-like cells differentiated in this manner exhibit surfactant processing and secretion capabilities, and long-term commitment to a mature AT2 phenotype when expanded in media optimized for primary AT2 culture. Comparing AT2-like cells differentiated with TGF{beta}-inhibition and BMP-activation to alternative differentiation approaches revealed improved specificity to the AT2 lineage and reduced off-target cell types. These findings reveal opposing roles for TGF{beta}- and BMP-signaling in AT2 differentiation and provide a new strategy to generate a therapeutically relevant cell type in vitro.

developmental biology↗

bioRxiv2022

Distinct airway progenitor cells drive epithelial heterogeneity in the developing human lung

Recent advances using single cell genomic approaches have identified new epithelial cell types and uncovered cellular heterogeneity in the murine and human lung (1). Here, using scRNA-seq and microscopy we identify and describe a secretory-like cell that is enriched in the small airways of the developing human lung and identified by the unique co-expression of SCGB3A2/SFTPB/CFTR. To place these cells in the hierarchy of airway development, we apply a single cell barcode-based lineage tracing method track the fate of SCGB3A2/SFTPB/CFTR cells during airway organoid differentiation in vitro (2). Lineage tracing revealed that these cells have distinct developmental potential from basal cells, giving rise predominantly to pulmonary neuroendocrine cells (PNECs) and a subset of multiciliated cells distinguished by high C6 and low MUC16 expression. We conclude that SCGB3A2/SFTPB/CFTR cells act as a progenitor cell contributing to the cellular diversity and heterogeneity in the developing human airway. SIGNIFICANCE STATEMENTThe current study identifies a novel secretory cell type that is present predominantly in the small airway of the developing human lung. These secretory cells are defined by co-expression of SCGB3A2/SFTPB/CFTR, and functional studies show that this cell gives rise to pulmonary neuroendocrine cells and a sub-population of multiciliated cells, thereby leading to cellular heterogeneity.

developmental biology↗

bioRxiv2022

Stable iPSC-derived NKX2-1+ Lung Bud Tip Progenitor Organoids Give Rise to Airway and Alveolar Cell Types

Bud tip progenitors (BTPs) in the developing lung give rise to all epithelial cell types found in the airways and alveoli. The current work aimed to develop an iPSC organoid model enriched with stable NKX2-1+ BTP-like cells. Building on prior work, we optimized a directed differentiation paradigm to generate spheroids with robust NKX2-1 expression. Spheroids were expanded into organoids that possessed NKX2-1+/CPM+ BTP-like cells, which increased in number over time. Single cell RNA-sequencing analysis revealed a high degree of transcriptional similarity between induced BTPs (iBTPs) and in vivo BTPs. Using FACS, iBTPs can be purified and expanded as induced bud tip organoids (iBTO), which maintain an enriched population of bud tip progenitors. When iBTOs are directed to differentiate into airway or alveolar cell types using well-established methods, they give rise to organoids composed of organized airway or alveolar epithelium, respectively. Collectively, iBTOs are transcriptionally and functionally similar to in vivo BTPs, providing an important model to study human lung development and differentiation. SUMMARY STATEMENTiPSC-derived lung bud tip progenitors emerge in organoid culture, can be isolated and expanded, are transcriptionally similar to primary bud tip progenitors, and can differentiate into airway or alveolar organoids.

developmental biology↗

bioRxiv2021

R-SPONDIN2+ Mesenchymal Cells Form the Bud Tip Progenitor Niche During Human Lung Development

Mammalian respiratory system development is regulated by complex reciprocal signaling events that take place between epithelial cells and the surrounding mesenchymal cells; however, mesenchymal heterogeneity and function in the developing human lung is poorly understood. We interrogated single cell RNA sequencing data from multiple human lung specimens and identified a mesenchymal cell population present during development that is highly enriched for expression of the WNT agonist R-SPONDIN2 (RSPO2), and we found that adjacent epithelial bud tip progenitors are enriched for the RSPO2 receptor LGR5. By carrying out functional experiments using organoid models, lung explant cultures, and FACS-isolated RSPO2+ mesenchyme, we show that RSPO2 is a critical niche cue that potentiates WNT signaling in human lung progenitors to maintain their multipotency.

developmental biology↗