Although single-cell RNA sequencing studies have begun to provide compendia of cell expression profiles
, it has been difficult to systematically identify and localize all molecular cell types in ...individual organs to create a full molecular cell atlas. Here, using droplet- and plate-based single-cell RNA sequencing of approximately 75,000 human cells across all lung tissue compartments and circulating blood, combined with a multi-pronged cell annotation approach, we create an extensive cell atlas of the human lung. We define the gene expression profiles and anatomical locations of 58 cell populations in the human lung, including 41 out of 45 previously known cell types and 14 previously unknown ones. This comprehensive molecular atlas identifies the biochemical functions of lung cells and the transcription factors and markers for making and monitoring them; defines the cell targets of circulating hormones and predicts local signalling interactions and immune cell homing; and identifies cell types that are directly affected by lung disease genes and respiratory viruses. By comparing human and mouse data, we identified 17 molecular cell types that have been gained or lost during lung evolution and others with substantially altered expression profiles, revealing extensive plasticity of cell types and cell-type-specific gene expression during organ evolution including expression switches between cell types. This atlas provides the molecular foundation for investigating how lung cell identities, functions and interactions are achieved in development and tissue engineering and altered in disease and evolution.
Elucidating the spectrum of epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) states in clinical samples promises insights on cancer progression and drug resistance. ...Using mass cytometry time-course analysis, we resolve lung cancer EMT states through TGFβ-treatment and identify, through TGFβ-withdrawal, a distinct MET state. We demonstrate significant differences between EMT and MET trajectories using a computational tool (TRACER) for reconstructing trajectories between cell states. In addition, we construct a lung cancer reference map of EMT and MET states referred to as the EMT-MET PHENOtypic STAte MaP (PHENOSTAMP). Using a neural net algorithm, we project clinical samples onto the EMT-MET PHENOSTAMP to characterize their phenotypic profile with single-cell resolution in terms of our in vitro EMT-MET analysis. In summary, we provide a framework to phenotypically characterize clinical samples in the context of in vitro EMT-MET findings which could help assess clinical relevance of EMT in cancer in future studies.
Volumetric muscle loss (VML) is associated with loss of skeletal muscle function, and current treatments show limited efficacy. Here we show that bioconstructs suffused with genetically-labelled ...muscle stem cells (MuSCs) and other muscle resident cells (MRCs) are effective to treat VML injuries in mice. Imaging of bioconstructs implanted in damaged muscles indicates MuSCs survival and growth, and ex vivo analyses show force restoration of treated muscles. Histological analysis highlights myofibre formation, neovascularisation, but insufficient innervation. Both innervation and in vivo force production are enhanced when implantation of bioconstructs is followed by an exercise regimen. Significant improvements are also observed when bioconstructs are used to treat chronic VML injury models. Finally, we demonstrate that bioconstructs made with human MuSCs and MRCs can generate functional muscle tissue in our VML model. These data suggest that stem cell-based therapies aimed to engineer tissue in vivo may be effective to treat acute and chronic VML.
Metabolic reprogramming of the tumor microenvironment is recognized as a cancer hallmark. To identify new molecular processes associated with tumor metabolism, we analyzed the transcriptome of bulk ...and flow-sorted human primary non-small cell lung cancer (NSCLC) together with
FDG-PET scans, which provide a clinical measure of glucose uptake. Tumors with higher glucose uptake were functionally enriched for molecular processes associated with invasion in adenocarcinoma and cell growth in squamous cell carcinoma (SCC). Next, we identified genes correlated to glucose uptake that were predominately overexpressed in a single cell-type comprising the tumor microenvironment. For SCC, most of these genes were expressed by malignant cells, whereas in adenocarcinoma, they were predominately expressed by stromal cells, particularly cancer-associated fibroblasts (CAF). Among these adenocarcinoma genes correlated to glucose uptake, we focused on glutamine-fructose-6-phosphate transaminase 2 (
), which codes for the glutamine-fructose-6-phosphate aminotransferase 2 (GFAT2), a rate-limiting enzyme of the hexosamine biosynthesis pathway (HBP), which is responsible for glycosylation.
was predictive of glucose uptake independent of GLUT1, the primary glucose transporter, and was prognostically significant at both gene and protein level. We confirmed that normal fibroblasts transformed to CAF-like cells, following TGFβ treatment, upregulated HBP genes, including
, with less change in genes driving glycolysis, pentose phosphate pathway, and TCA cycle. Our work provides new evidence of histology-specific tumor stromal properties associated with glucose uptake in NSCLC and identifies
as a critical regulator of tumor metabolic reprogramming in adenocarcinoma.
These findings implicate the hexosamine biosynthesis pathway as a potential new therapeutic target in lung adenocarcinoma.
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Mechanical ventilation (MV) is a life-saving measure for those incapable of adequately ventilating or oxygenating without assistance. Unfortunately, even brief periods of MV result in diaphragm ...weakness (i.e., ventilator-induced diaphragm dysfunction VIDD) that may render it difficult to wean the ventilator. Prolonged MV is associated with cascading complications and is a strong risk factor for death. Thus, prevention of VIDD may have a dramatic impact on mortality rates. Here, we summarize the current understanding of the pathogenic events underlying VIDD. Numerous alterations have been proven important in both human and animal MV diaphragm. These include protein degradation via the ubiquitin proteasome system, autophagy, apoptosis, and calpain activity-all causing diaphragm muscle fiber atrophy, altered energy supply via compromised oxidative phosphorylation and upregulation of glycolysis, and also mitochondrial dysfunction and oxidative stress. Mitochondrial oxidative stress in fact appears to be a central factor in each of these events. Recent studies by our group and others indicate that mitochondrial function is modulated by several signaling molecules, including Smad3, signal transducer and activator of transcription 3, and FoxO. MV rapidly activates Smad3 and signal transducer and activator of transcription 3, which upregulate mitochondrial oxidative stress. Additional roles may be played by angiotensin II and leaky ryanodine receptors causing elevated calcium levels. We present, here, a hypothetical scaffold for understanding the molecular pathogenesis of VIDD, which links together these elements. These pathways harbor several drug targets that could soon move toward testing in clinical trials. We hope that this review will shape a short list of the most promising candidates.
Talcum has been used in pleurodesis for more than 8 decades. Despite a wealth of research, controversy remains over the optimal sclerosant for pneumothorax and pleural effusions. Talc’s historical ...primacy has been challenged because of its potential for pulmonary toxicity, possible carcinogenicity, and recent concerns surrounding availability and legal liability, thus making this an ideal time for a review.
This systematic review of the talc literature, focused on publications after the year 2000, evaluated mechanism of action, efficacy, side effect profile, and alternative sclerosants; included is an overview of current socioeconomic and legal controversies.
The data support talc as the most effective agent for pleurodesis. There is evidence to suggest that mean particle size has a direct relationship with the side effect profile and that significant hypoxemic events after talc administration are exceedingly rare when using available graded talc preparations. Concerns regarding the development of malignant diseases after topical talc application remain incompletely resolved but appear related to cosmetic powder preparations that were contaminated with asbestos. Purified talc in the pleural space has not been implicated. Recent difficulties accessing commercial talc preparations have been solved. Although safe and effective talc alternatives do exist, these agents are not as well studied.
Talc pleurodesis with modern, purified, graded talc preparations is safe and highly effective. Talc is an inexpensive and accessible option that remains appropriate for pleurodesis despite existing controversies.
Adult skeletal muscle stem cells, or satellite cells (SCs), regenerate functional muscle following transplantation into injured or diseased tissue. To gain insight into human SC (huSC) biology, we ...analyzed transcriptome dynamics by RNA sequencing of prospectively isolated quiescent and activated huSCs. This analysis indicated that huSCs differentiate and lose proliferative potential when maintained in high-mitogen conditions ex vivo. Further analysis of gene expression revealed that p38 MAPK acts in a transcriptional network underlying huSC self-renewal. Activation of p38 signaling correlated with huSC differentiation, while inhibition of p38 reversibly prevented differentiation, enabling expansion of huSCs. When transplanted, expanded huSCs differentiated to generate chimeric muscle and engrafted as SCs in the sublaminar niche with a greater frequency than freshly isolated cells or cells cultured without p38 inhibition. These studies indicate characteristics of the huSC transcriptome that promote expansion ex vivo to allow enhanced functional engraftment of a defined population of self-renewing huSCs.
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•Prospective isolation of highly pure huSCs from diverse muscles•RNA sequencing resource for studying the huSC transcriptome•Core transcription factor regulatory network of huSC differentiation•Expanded huSCs that are genetically manipulable and self-renew in vivo
In this article, Rando and colleagues purify and study human skeletal muscle stem cells using RNA sequencing and cell transplantation. They show that p38 MAPK signaling is important for human muscle stem cell differentiation and that pharmacologic inhibition of p38 enables expansion of muscle stem cells capable of self-renewing after transplantation.
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