Adoptive Transfer of iTregs for FVIII Tolerance

One million pltLys-iTreg or purTGFβ-iTreg cells were adoptively transferred by IV injection into F8^null/B6 mice followed by rhF8 (Xyntha; Pfizer Inc., New York, NY) immunization weekly at 50 U/kg per week IV for a total of 4 weeks. One week after the last immunization, blood samples were collected. The titers of anti-FVIII inhibitory antibodies (inhibitors) and total anti-FVIII antibodies were determined by Bethesda assay and enzyme-linked immunosorbent assay (ELISA), respectively, as described in our previous report.^{16 (link),19 (link)} Six days after the iTreg cell transfer, CD4⁺ T cells from peripheral blood were analyzed by flow cytometry for enhanced green fluorescent protein (EGFP) expression to assess the recovery of the infused iTregs. CD45.1 and CD45.2 were used as congenic markers.

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Haribhai D., Luo X., Chen J., Jia S., Shi L., Schroeder J.A., Weiler H., Aster R.H., Hessner M.J., Hu J., Williams C.B, & Shi Q. (2016). TGF-β1 along with other platelet contents augments Treg cells to suppress anti-FVIII immune responses in hemophilia A mice. Blood Advances, 1(2), 139-151.

Publication 2016

Xyntha

Anti antibodies Assay Blood Cell Enhanced green fluorescent protein Enzyme linked immunosorbent assay Flow cytometry Immunization Inhibitors Inhibitory Mice Peripheral blood cells Xyntha

Corresponding Organization : Versiti Blood Center of Wisconsin

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Variable analysis

independent variables

Adoptive transfer of one million pltLys-iTreg or purTGFβ-iTreg cells by IV injection into F8^null/B6 mice
Weekly rhF8 (Xyntha; Pfizer Inc., New York, NY) immunization at 50 U/kg per week IV for a total of 4 weeks

dependent variables

Titers of anti-FVIII inhibitory antibodies (inhibitors) determined by Bethesda assay
Titers of total anti-FVIII antibodies determined by enzyme-linked immunosorbent assay (ELISA)
Recovery of the infused iTregs assessed by flow cytometry for enhanced green fluorescent protein (EGFP) expression in CD4⁺ T cells from peripheral blood

control variables

F8^null/B6 mouse strain
CD45.1 and CD45.2 congenic markers used to identify the infused iTregs

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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