Boudreau, D. K. | bioRxiv Science

bioRxiv2025

Cell-Specific Transcriptomic and Mito-Nuclear Imbalance in Lungs Under Intermittent Hypoxia in Adult Male Mice.

Obstructive sleep apnea and its characteristic intermittent hypoxia (IH) are widely recognized as significant contributors to various pulmonary diseases, including asthma, pulmonary arterial hypertension, fibrosis, and chronic obstructive pulmonary disease. While single-cell RNA sequencing (scRNA-seq) has provided valuable insights into cell-type-specific responses to IH, previous studies have primarily focused on post-hypoxic recovery states, leaving immediate molecular responses during active IH exposure unexplored. To address this critical knowledge gap, we investigated real-time transcriptional responses to IH at single-cell resolution in lung tissue using male mice (n=3/group) exposed to either normoxia or IH (30 cycles/h, nadir 6% O2, 12 h/day) for 14 days, with tissue collection during active IH exposure. Our analysis revealed pronounced cell-type-specific transcriptional reprogramming, particularly in airway smooth muscle cells (ASMC), arterial endothelial cells (AEC), and lymphatic endothelial cells (LEC). These changes were characterized by enrichment in pathways related to epithelial-to-mesenchymal transition (ASMC, LEC), myogenesis (ASMC), and antioxidant defenses (AEC, LEC). Most cell types demonstrated substantial upregulation of genes encoding mitochondrial complex I-IV proteins and TCA cycle enzymes accompanied by a decreased expression of genes encoded by mitochondrial DNA that was markedly present in LEC, AEC, and cells of the alveolar-capillary unit, revealing a mito-nuclear imbalance. These findings provide novel insights into the immediate cellular responses to IH, showing previously uncharacterized metabolic reorganization that may underlie the development of IH-related pulmonary complications. This improved understanding of early molecular events during active IH exposure advances our knowledge of sleep apnea-related lung pathologies and may inform future therapeutic strategies.

physiology↗

bioRxiv2024

Single-cell and single-nucleus RNA-sequencing from paired normal-adenocarcinoma lung samples provides both common and discordant biological insights.

Whether single-cell RNA-sequencing (scRNA-seq) captures the same biological information as single-nucleus RNA-sequencing (snRNA-seq) remains uncertain and likely to be context-dependent. Herein, a head-to-head comparison was performed in matched normal-adenocarcinoma human lung samples to assess biological insights derived from scRNA-seq versus snRNA-seq and better understand the cellular transition that occurs from normal to tumoral tissue. Here, the transcriptome of 160,621 cells/nuclei was obtained. In non-tumor lung, cell type proportions varied widely between scRNA-seq and snRNA-seq with a predominance of immune cells in the former (81.5%) and epithelial cells (69.9%) in the later. Similar results were observed in adenocarcinomas, in addition to an overall increase in cell type heterogeneity and a greater prevalence of copy number variants in cells of epithelial origin, which suggests malignant assignment. The cell type transition that occurs from normal lung tissue to adenocarcinoma was not always concordant whether cells or nuclei were examined. As expected, large differential expression of the whole-cell and nuclear transcriptome was observed, but cell-type specific changes of paired normal and tumor lung samples revealed a set of common genes in the cells and nuclei involved in cancer-related pathways. In addition, we showed that the ligand-receptor interactome landscape of lung adenocarcinoma was largely different whether cells or nuclei were evaluated. Immune cell depletion in fresh specimens partly mitigated the difference in cell type composition observed between cells and nuclei. However, the extra manipulations affected cell viability and amplified the transcriptional signatures associated with stress responses. In conclusion, research applications focussing on mapping the immune landscape of lung adenocarcinoma benefit from scRNA-seq in fresh samples, whereas snRNA-seq of frozen samples provide a low-cost alternative to profile more epithelial and cancer cells, and yield cell type proportions that more closely match tissue content. Author SummarySingle-cell transcriptomic datasets provide unprecedented opportunities to disentangle the complex tissue microenvironment and cellular origin of cancer. Data are scarce regarding the pros and cons of single-cell RNA sequencing (scRNA-seq) of freshly explanted human tissues over single-nuclei sequencing (snRNA-seq) from the same archived frozen tissues. Lung adenocarcinoma represents a medically valuable case study to compare the biological signal recovered through cells and nuclei sequencing. Here, we sequenced the transcriptome of 160,621 cells/nuclei in paired normal-adenocarcinoma lung samples. Cell type proportions varied widely between scRNA-seq and snRNA-seq with a predominance of immune cells in the former and epithelial cells in the later. Adenocarcinomas were characterized by an increase in cell type heterogeneity and a greater prevalence of malignant epithelial cells in both scRNA-seq and snRNA-seq. The cellular and gene expression transition that occur from normal lung to adenocarcinoma showed common and discordant biological insights whether cells or nuclei were examined. Research applications focussing on mapping the immune landscape of lung cancer benefit from scRNA-seq in fresh samples, whereas snRNA-seq of the same frozen samples provide a low-cost and more flexible alternative to profile more epithelial and cancer cells, and yield cell type proportions that more closely match tissue content.

cancer biology↗