Increased Endothelial Colony-Forming Cells (ECFC) and Endothelial Microparticles (EMP) and Their Interaction in Thrombotic Thrombocytopenic Purpura (TTP).
Authors: Jimenez Joaquin J. , Jy Wenche , Mauro Lucia M. , Bidot Loreta , Salgueiro Luis M. , Andrews David , Shariatmadar Sherry , Horstman Lawrence L. , Fontana Vincenzo , Ahn Yeon-Soong 
Published: April 2020

BACKGROUND. Endothelial damage is a prominent feature of TTP, evidenced by elevated EMP levels during acute episodes of TTP. However, endothelial repair in TTP remains to be investigated. ECFC are key players in endothelial regeneration, and their assay in blood has been employed to monitor endothelial remodeling. We investigated circulating ECFC in TTP and their relationship with EMP.

METHODS. Six patients with acquired TTP were studied. All were treated with therapeutic plasma exchange (TPE). All but one underwent complete remission (CR). The one who failed to achieve CR with TPE suffered multiple strokes and developed chronic renal failure. We measured the following in acute phase and remission, as well as pre- and post-TPE:

EMP defined by CD31+/CD42b- (EMP31) or CD62E+ (EMP62) by flow cytometry; ECFC in blood by the method of Ingram et al [Blood 104:2752, 2004]; ADAMTS13 activity in blood as well as in supernatants of ECFC-derived EC by the FRETS-vWF73 fluorogenic method. In addition, we investigated effects of microparticles (MP) from TTP plasma on normal ECFC.

In separate experiments, EC cultured from TTP patients were assayed for surface expression of CD106 and CD54, and EMP released in response to TNF-α.

RESULTS.

In the acute phase, EMP were elevated: EMP31 (1.89 ± 0.65 × 106 counts/mL vs. 0.28 ±0.1 for controls, p=0.001) and EMP62 (5.7 ±1.3 × 106 counts/mL vs. 0.40 ±0.12 for controls, p&lt;0.001). After the first TPE, EMP fell significantly. In acute episodes of TTP, ECFC colonies were significantly increased in 5 responders (4.1 ±0.5 vs. 1.0 ±0.03 for controls, p&lt;0.01). However, no elevation in ECFC was observed in the non-responder. Upon remission, ECFC colonies returned to normal in responders. ADAMTS13 activity was &lt;10% in acute phase and rose after TPE. A positive correlation was found between initial EMP31 or EMP62 levels and ECFC (R=0.60 and R=0.71, respectively; p&lt;0.001).

To test whether this correlation might be due to ECFC-EMP interaction, we studied the MP fraction from the 5 responders on ECFC. A significant increase in colony counts was observed upon treatment with 1 × 108 MP/mL (1.2 ±0.04 ECFC colonies vs. 2.0 ±0.5 MP-treated, p=0.01). Thus, MP apparently stimulate ECFC colony formation. We have also tested the functional activities of ECFC from TTP patients. No significant difference was observed in ADAMTS13 activity secreted by EC from TTP compared to control or remission EC. In addition, EC cultures derived from ECFC colonies from TTP patients did not differ from controls in response to TNF-α stimulation, neither in EMP released or expression of surface markers.

CONCLUSIONS. TPE decreases plasma EMP in all responders. The benefit of TPE may be attributable in part to removal of EMP and other procoagulant MP. The increased ECFC in acute TTP, demonstrated for the first time in this study, apparently reflects enhanced endothelial repair in the face of injurious processes among responders. Failure of this regeneration may entail irreversible damage, as seen in the non-responder. Finally, EMP or other MP apparently stimulate ECFC and may play a role in endothelial repair through interaction with ECFC.