Autophagy in endothelial cells regulates their haematopoiesis-supporting ability

Abstract

Background: Endothelial cells (ECs) function as an instructive platform to support haematopoietic stem cell (HSC) homeostasis. Our recent studies found that impaired bone marrow (BM) ECs are responsible for the defective haematopoiesis in patients with poor graft function (PGF), which is characterised by pancytopenia post-allotransplant. Although activated autophagy was reported to benefit ECs, whether EC autophagy plays a critical role in supporting HSCs and its effect on PGF patients post-allotransplant remain unclear. Methods: To evaluate whether the autophagy status of ECs modulates their ability to support haematopoiesis, human umbilical vein endothelial cells (HUVECs) and primary BM ECs derived from healthy donors were subjected to knockdown or overexpression of Beclin-1 (an autophagy-related protein). Moreover, BM ECs derived from PGF patients were studied. Findings: Beclin-1 knockdown significantly reduced the haematopoiesis-supporting ability of ECs by suppressing autophagy, which could be restored by activating autophagy via Beclin-1 upregulation. Moreover, autophagy positively regulated haematopoiesis-related genes in HUVECs. Subsequently, a prospective casecontrol study demonstrated that defective autophagy reduced Beclin-1 expression and the colony-forming unit (CFU) plating efficiency in BM ECs from PGF patients compared to matched patients with good graft function. Rapamycin, an autophagy activator, quantitatively and functionally improved BM ECs from PGF patients in vitro and enhanced their ability to support HSCs by activating the Beclin-1 pathway. Interpretation: Our results suggest that the autophagy status of ECs modulates their ability to support haematopoiesis by regulating the Beclin-1 pathway. Defective autophagy in BM ECs may be involved in the pathogenesis of PGF post-allotransplant. Rapamycin provides a promising therapeutic approach for PGF patients. Funding: Please see funding sources.

Oxidative Stress Triggers Defective Autophagy in Endothelial Cells: Role in Atherothrombosis Development

“…Moreover, in ECs, the autophagic process significantly supports haematopoietic stem cell homeostasis. Indeed, the impairment of bone marrow ECs is prevented by the modulation of Beclin-1 signaling, which activates autophagy in patients with poor graft function [ 100 ].…”

Improved function and balance in T cell modulation by endothelial cells in young people

“…The following multicolour flow cytometry panel were applied to quantify ECs in BM: anti-CD34-PE/Cy7, anti-CD309-PE, and anti-CD133-APC (BD Biosciences, CA, USA). ECs were identified as CD34 + CD309 + CD133 + , and the relative EC frequency was expressed as a fraction of BMMNCs [23][24][25][26][27]. Samples were collected on a BD LSRFortessa flow cytometer and the compensation adjustion as well as data analysis were accomplished via a BD LSRFortessa software.…”

“…To examine EC ROS level, 10 μmol/l DCFH-DA (Beyotime, Beijing, China) was added into the BMMNC suspension that stained with EC markers at 37°C for 15 minutes [23][24][25][26][27]. EC ROS levels were examined by flow cytometry and expressed as the mean fluorescence intensity (MFI).…”

“…As previously reported [23][24][25][26][27], BMMNCs were extracted and seeded to the cell culture plates (Corning, NY, USA) pre-coated with fibronectin (Sigma, MO, USA). EGM-2-BulletKit (Lonza, MD, USA) medium with extra 10% foetal bovine serum (FBS) (Gibco, MA, USA) was added to the cell plates and was renewed on the fourth day to supplement nutrition for cells.…”

Autophagy in the Regulation of Tissue Differentiation and Homeostasis

“…Endothelial cells are also essential elements of the hematopoietic SC (HSC) niche by providing critical signals to support blood cell production in the bone marrow ( Mendelson and Frenette, 2014 ). Again, autophagy has been proposed as a key regulator of this process because beclin-1 knockdown reduces the hematopoiesis-supporting ability of ECs that can be restored by beclin-1 upregulation ( Lyu et al, 2020 ). Notably, a prospective case-control study illustrates that defective autophagy of ECs in the HSC niche may be involved in the post-allotransplant pancytopenia characteristic of poor graft function (PGF), suggesting the possibility of treating PGF patients with drugs able to promote autophagy (for example, rapamycin).…”