Clinically compatible advances in blood-derived endothelial progenitor cell isolation and reprogramming for translational applications

The promise of using induced pluripotent stem cells (iPSCs) for cellular therapies has been hampered by the lack of easily isolatable and well characterized source cells whose genomes have undergone minimal changes during their processing. Blood-derived late-outgrowth endothelial progenitor cells (EPCs) are used for disease modeling and have potential therapeutic uses including cell transplantation and the translation of induced pluripotent stem cell (iPSC) derivatives. However, the current isolation of EPCs has been inconsistent and requires at least 40−80 mL of blood, limiting their wider use. In addition, previous EPC reprogramming methods precluded the translation of EPC-derived iPSCs to the clinic. Here a series of clinically-compatible advances in the isolation and reprogramming of EPCs is presented, including a reduction of blood sampling volumes to 10 mL and use of highly efficient RNA-based reprogramming methods together with autologous human serum, resulting in clinically relevant iPSCs carrying minimal copy number variations (CNVs) compared to their parent line

Autophagy contributes to BMP type 2 receptor degradation and development of pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) is characterised by an increase in mean pulmonary arterial pressure which almost invariably leads to right heart failure and premature death. More than 70% of familial PAH and 20% of idiopathic PAH patients carry heterozygous mutations in the bone morphogenetic protein (BMP) type 2 receptor (BMPR2). However, the incomplete penetrance of BMPR2 mutations suggests that other genetic and environmental factors contribute to the disease. In the current study, we investigate the contribution of autophagy in the degradation of BMPR2 in pulmonary vascular cells. We demonstrate that endogenous BMPR2 is degraded through the lysosome in primary human pulmonary artery endothelial (PAECs) and smooth muscle cells (PASMCs): two cell types that play a key role in the pathology of the disease. By means of an elegant HaloTag system, we show that a block in lysosomal degradation leads to increased levels of BMPR2 at the plasma membrane. In addition, pharmacological or genetic manipulations of autophagy allow us to conclude that autophagy activation contributes to BMPR2 degradation. It has to be further investigated whether the role of autophagy in the degradation of BMPR2 is direct or through the modulation of the endocytic pathway. Interestingly, using an iPSC-derived endothelial cell model, our findings indicate that BMPR2 heterozygosity alone is sufficient to cause an increased autophagic flux. Besides BMPR2 heterozygosity, pro-inflammatory cytokines also contribute to an augmented autophagy in lung vascular cells. Furthermore, we demonstrate an increase in microtubule-associated protein 1 light chain 3 beta (MAP1LC3B) levels in lung sections from PAH induced in rats. Accordingly, pulmonary microvascular endothelial cells (MVECs) from end-stage idiopathic PAH patients present an elevated autophagic flux. Our findings support a model in which an increased autophagic flux in PAH patients contributes to a greater decrease in BMPR2 levels. Altogether, this study sheds light on the basic mechanisms of BMPR2 degradation and highlights a crucial role for autophagy in PAH

Search for gravitational waves from Scorpius X-1 with a hidden Markov model in O3 LIGO data

Results are presented for a semicoherent search for continuous gravitational waves from the low-mass x-ray binary Scorpius X-1, using a hidden Markov model (HMM) to allow for spin wandering. This search improves on previous HMM-based searches of Laser Interferometer Gravitational-Wave Observatory data by including the orbital period in the search template grid, and by analyzing data from the latest (third) observing run. In the frequency range searched, from 60 to 500 Hz, we find no evidence of gravitational radiation. This is the most sensitive search for Scorpius X-1 using a HMM to date. For the most sensitive subband, starting at 256.06 Hz, we report an upper limit on gravitational wave strain (at 95% confidence) of h 95 % 0 = 6.16 × 10 − 26 , assuming the orbital inclination angle takes its electromagnetically restricted value ι = 4 4 ° . The upper limits on gravitational wave strain reported here are on average a factor of ∼ 3 lower than in the second observing run HMM search. This is the first Scorpius X-1 HMM search with upper limits that reach below the indirect torque-balance limit for certain subbands, assuming ι = 4 4 °