In this issue, Xia et al. explore the role of delta-like 4 (DLL4), an endothelial cell Notch ligand, in the regulation of lung microvascularization and distal lung morphogenesis. Using lung mapping approaches in mouse and human cadaveric samples, as well functional studies in Dll4+/– mice, they show that DLL4 is required for pulmonary microvascularization and alveolarization during lung development. The cover image shows a postnatal day 6 lung sample from a Dll4+/lacZ reporter mouse stained with X-gal (blue) and platelet endothelial cell adhesion molecule (PECAM, brown) and counterstained with Nuclear Fast Red.
Skeletal muscle can regenerate from muscle stem cells and their myogenic precursor cell progeny, myoblasts. However, precise gene editing in human muscle stem cells for autologous cell replacement therapies of untreatable genetic muscle diseases has not yet been reported. Loss-of-function mutations in SGCA, encoding α-sarcoglycan, cause limb-girdle muscular dystrophy 2D/R3, an early onset, severe and rapidly progressive form of muscular dystrophy affecting equally girls and boys. Patients suffer from muscle degeneration and atrophy affecting the limbs, respiratory muscles, and the heart. We isolated human muscle stem cells from two donors with the common SGCA c.157G>A mutation affecting the last coding nucleotide of exon 2. We found that c.157G>A is an exonic splicing mutation that induces skipping of two co-regulated exons. Using adenine base editing, we corrected the mutation in the cells from both donors with >90% efficiency, thereby rescuing the splicing defect and α-sarcoglycan expression. Base edited patient cells regenerated muscle and contributed to the Pax7 positive satellite cell compartment in vivo in mouse xenografts. We hereby provide the first evidence that autologous gene repaired human muscle stem cells can be harnessed for cell replacement therapies of muscular dystrophies.
Helena Escobar, Anne Krause, Sandra Keiper, Janine Kieshauer, Stefanie Müthel, Manuel García de Paredes, Eric Metzler, Ralf Kühn, Florian Heyd, Simone Spuler
Bariatric surgery is the most effective method for weight loss in morbid obesity. There is significant individual variability in the weight loss outcomes, yet factors leading to postoperative weight loss or weight regain remain elusive. Alterations in the µ-opioid receptor (MOR) and dopamine D2 receptor (D2R) systems are associated with obesity and appetite control, and the magnitude of initial brain receptor system perturbation may predict long-term surgical weight loss outcomes. We tested this hypothesis by studying 19 morbidly obese women (mean BMI 40) scheduled to undergo bariatric surgery. We measured their preoperative MOR and D2R availabilities using positron emission tomography (PET) with [11C]carfentanil and [11C]raclopride, respectively, and then assessed their weight development association with regional MOR and D2R availabilities at 24-month follow-up. MOR availability in the amygdala consistently predicted weight development throughout the follow-up period, but no associations were found for D2R. This is the first study to demonstrate that neuroreceptor markers prior to bariatric surgery are associated with the postoperative weight loss. Postoperative weight regain may derive from dysfunction in the opioid system, and weight loss outcomes after bariatric surgery may be partially predicted based on preoperative brain receptor availability opening up new potential for treatment possibilities.
Henry K. Karlsson, Lauri Tuominen, Semi Helin, Paulina Salminen, Pirjo Nuutila, Lauri Nummenmaa
BACKGROUND. Idiopathic intracranial hypertension (IIH) is a condition predominantly affecting obese women of reproductive age. Recent evidence suggests that IIH is a disease of metabolic dysregulation, androgen excess and an increased risk of cardiovascular morbidity. Here we evaluate systemic and adipose specific metabolic determinants of the IIH phenotype. METHODS. In fasted, matched IIH (N=97) and control (N=43) patients, we assessed: glucose and insulin homeostasis and leptin levels. Body composition was assessed along with an interrogation of adipose tissue function via nuclear magnetic resonance metabolomics and RNA sequencing in paired omental and subcutaneous biopsies in a case control study. RESULTS. We demonstrate an insulin and leptin resistant phenotype in IIH in excess to that driven by obesity. Adiposity in IIH is preferentially centripetal and is associated with increased disease activity and insulin resistance. IIH adipocytes appear transcriptionally and metabolically primed towards depot-specific lipogenesis. CONCLUSIONS. These data show that IIH is a metabolic disorder in which adipose tissue dysfunction is a feature of the disease. Managing IIH as a metabolic disease could reduce disease morbidity and improving cardiovascular outcomes. FUNDING. This study was supported by the National Institute of Health Research UK (NIHR-CS-011-028), the Medical Research Council UK (MR/K015184/1) and the Midlands Neuroscience Teaching and Research Fund.
Connar S.J. Westgate, Hannah F. Botfield, Zerin Alimajstorovic, Andreas Yiangou, Mark Walsh, Gabrielle Smith, Rishi Singhal, James L. Mitchell, Olivia Grech, Keira A. Markey, Daniel Hebenstreit, Daniel A. Tennant, Jeremy W. Tomlinson, Susan P. Mollan, Christian Ludwig, Ildem Akerman, Gareth G. Lavery, Alexandra J. Sinclair
Retinoic acid (RA) signaling is essential for enteric nervous system (ENS) development since vitamin A deficiency or mutations in RA signaling profoundly reduce bowel colonization by ENS precursors. These RA effects could occur because of RA activity within the ENS lineage or via RA activity in other cell types. To define cell-autonomous roles for retinoid signaling within the ENS lineage at distinct developmental time points, we activated a potent floxed dominant-negative RA receptor α (RarαDN) in the ENS using diverse CRE recombinase-expressing mouse lines. This strategy enabled us to block RA signaling at pre-migratory, migratory, and post-migratory stages for ENS precursors. We found that cell-autonomous loss of retinoic acid receptor (RAR) signaling dramatically affects ENS development. CRE activation of RarαDN expression at pre-migratory or migratory stages caused severe intestinal aganglionosis, but at later stages, RarαDN induced a broad range of phenotypes including hypoganglionosis, submucosal plexus loss, and abnormal neural differentiation. RNA-sequencing highlighted distinct RA-regulated gene sets at different developmental stages. These studies show complicated context-dependent RA-mediated regulation of ENS development.
Tao Gao, Elizabeth C. Wright-Jin, Rajarshi Sengupta, Jessica B. Anderson, Robert O. Heuckeroth
Transitions between cell fates commonly occur in development and disease. However, reversing an unwanted cell transition in order to treat disease remains an unexplored area. Here, we report a successful process of guiding ill-fated transitions toward normalization in vascular calcification. Vascular calcification is a severe complication that increases all-cause mortality of cardiovascular disease but lacks medical therapy. The vascular endothelium is a contributor of osteoprogenitor cells to vascular calcification through endothelial-mesenchymal transitions, in which endothelial cells (ECs) gain plasticity and ability to differentiate into osteoblast-like cells. We created a high throughput screening and identified SB216763, an inhibitor of glycogen synthase kinase 3 (GSK3), as an inducer of osteoblastic-endothelial transition. We demonstrated that SB216763 limits osteogenic differentiation in ECs at an early stage of vascular calcification. Lineage tracing showed that SB216763 redirected osteoblast-like cells to the endothelial lineage and reduced late-stage calcification. We also find that deletion of GSK3beta in osteoblasts recapitulated osteoblastic-endothelial transition and reduced vascular calcification. Overall, inhibition of GSK3beta promoted the transition of cells with osteoblastic characteristic to endothelial differentiation thereby ameliorating vascular calcification.
Jiayi Yao, Xiuju Wu, Xiaojing Qiao, Daoqin Zhang, Li Zhang, Jocelyn A. Ma, Xinjiang Cai, Kristina I. Boström, Yucheng Yao
JCI This Month is a digest of the research, reviews, and other features published each month.