Pluronic-F127/Platelet Microvesicles nanocomplex delivers stem cells in high doses to the bone marrow and confers post-irradiation survival

Vikas Chander,Gurudutta Gangenahalli

doi:10.1038/s41598-019-57057-8

Abstract

Platelet microvesicles (pMVs) are submicron-sized heterogeneous vesicles released upon activation and contain several membrane receptors and proteins (CD41, CD61, CD62, CXCR4, PAR-1, etc.). We have revealed their ability to adhere to the triblock copolymer pluronic-F127 (PF127) and form a platelet microvesicular nanocloud which has the potential to enhance the transvascular migration of hematopoietic stem cells across the sinusoidal endothelium to the bone marrow. Besides, the pMVs nanoclouds bestow survival benefits when present on the cells used for infusion, particularly with PF127-stabilized with chitosan-alginate (PF127-CA HSCs). The vesicles were found to be firmly associated with PF127 in the nanocloud, which was detected by confocal laser scanning microscopy. The abrogation of CXCR4/SDF-1 axis regulating the transmigration of the cells by antagonist AMD3100 revealed that the enriched CXCR4 receptors on pMVs robustize the transmigration of the infused cells. The homing of the cells led to effective engraftment and faster regeneration of the critical blood lineages, which elicited 100% survival of the mice receiving lethal doses of radiation. The Human Long-Term Culture Initiating Cells (LTC-ICs), Severe Combined Immunodeficient (SCID) - Repopulating Cells (SRCs) and Colony Forming Cells (CFCs) responsible for the regeneration, but present in extremely low numbers in the infused cell dose, have enabled the cells to reach the bone marrow in high numbers. This potential of the PF127 to sequester the pMVs and its application to achieve over 10-fold delivery of HSCs across the trans-endothelial checkpoint has so far not been reported. Thus, this mechanistic innovation is a potential post-exposure life-saving regimen capable of circumventing the irreparable damage to the bone marrow caused by lethal doses of radiation.

Highlights

Pluronics, known as poloxamers, are amphiphilic triblock copolymers consisting of hydrophobic polyoxypropylene (PPO) and hydrophilic polyoxyethylene (PEO) domains
It is established that increased transvascular migration (TVM) enables atleast a few of the Severe Combined Immunodeficient (SCID) Repopulating Cells (SRC), Long Term Culture Initiating Cells (LTC-IC), and Colony Forming Cells (CFC) that are solely essential for regeneration, enter the bone marrow, restore the number of platelets and neutrophils to threshold levels which counter the thrombocytopenia and neutropenia, and confer survival
After establishing the stable presence of the shield for up to 8 hrs (Fig. 2(e) and Supplementary Fig. 2) and thereby completely preventing the binding of the cytokines to the cells (Supplementary Fig. 3), the PF127 layer was grafted at a non-toxic concentration (2 mg/ml), (Supplementary Fig. 4(a)) which was confirmed by confocal laser scanning microscopy (CLSM) (Fig. 3(a,b))

Summary

Introduction

Known as poloxamers, are amphiphilic triblock copolymers consisting of hydrophobic polyoxypropylene (PPO) and hydrophilic polyoxyethylene (PEO) domains. The mechanistic behavior of ensemble pMVs in the form of natural aggregates is similar to that of platelets and regulates different localized mechanisms, across the vascular endothelium, which induces certain pathophysiological responses such as homeostasis, thrombosis, atherosclerosis, and inflammation[14,15,16,17] When these pMVs are anchored to CXCR4 and other rolling and adhesion integrins, they tend to be responsive to SDF1 (CXCL12), a gradient (CXCR4/CXCL12 axis) that is consistently present only across the bone marrow due to secretions from the sinusoidal endothelium. We explored the likelihood of ensemble pMVs when densely made to present as platelet dust ( known as nanocloud) around the haematopoietic stem/progenitor cells (HSCs/HSPCs) We achieved this by grafting the triblock copolymer PF127 on a chitosan-alginate stabilized template over the cell surface to mediate the TVM of infused HSCs into the bone marrow.

Methods

Results

Conclusion