Beta 2-Glycoprotein I binds factor XI and inhibits its activation by thrombin and factor XIIa: loss of inhibition by clipped beta 2-glycoprotein I

Abstract

Activation of factor XI (FXI) by thrombin in vivo plays a role in coagulation by providing an important positive feedback mechanism for additional thrombin generation. FXI is activated in vitro by thrombin, or FXIIa in the presence of dextran sulfate. In this report, we investigated the effect of beta(2)-glycoprotein I (beta(2)GPI) on the activation of FXI. beta(2)GPI bound FXI in vitro and inhibited its activation to FXIa by thrombin and FXIIa. The affinity of the interaction between beta(2)GPI and FXI was equivalent to the interaction between FXI and high molecular weight kininogen. Inhibition of FXI activation occurred with lower concentrations of beta(2)GPI than found in human plasma. Proteolytic clipping of beta(2)GPI by plasmin abolished its inhibition of FXI activation. The results suggest a mechanism of regulation whereby physiological concentrations of beta(2)GPI may attenuate thrombin generation in vivo by inhibition of FXI activation. Plasmin cleavage of beta(2)GPI provides a negative feedback that counteracts its inhibition of FXI activation.

Fig. 1.

Binding of ¹²⁵I-FXI to rhβ₂GPI and nβ₂GPI. A constant concentration of ¹²⁵I-FXI was incubated in microtiter wells that were coated with serial dilutions of rhβ₂GPI (▴), nβ₂GPI (▾), or BSA (▪). The amount of ¹²⁵I-FXI binding to immobilized proteins was determined as described in Methods.

Fig. 2.

Inhibitory effects of rhβ₂GPI and nβ₂GPI on binding of ¹²⁵I-FXI to rhβ₂GPI. A constant concentration of ¹²⁵I-FXI was incubated with serial dilutions of rhβ₂GPI (▴), nβ₂GPI (▾), or BSA (▪) in wells of microtiter plates coated with rhβ₂GPI. The number of cpm bound were converted to percentage of total binding by dividing by the cpm bound in the absence of β₂GPI competitors.

Fig. 3.

β₂GPI dose-dependent inhibitory effect on the activation of FXI by thrombin and FXIIa in the presence of DS. Data are expressed as percentage of FXIa generated in the absence of inhibitor. ▪, Thrombin; □, FXIIa as activator.

Fig. 4.

Binding of ¹²⁵I-FXI to nβ₂GPI, cnβ₂GPI, and BSA (A), and activation of FXI (B) by thrombin in the presence of nβ₂GPI and cnβ₂GPI. The amount of ¹²⁵I-FXI bound and FXIa generated are expressed in pmol.

Fig. 5.

Activation of ¹²⁵I-FXI in β₂GPI-deficient human plasma. β₂GPI-deficient human plasma was reconstituted by the addition of rhβ₂GPI (▪) or BSA (□), and ¹²⁵I-FXI was added. (A) Autoradiograph. This is a representative example of three different experiments. (B) FXIa generated expressed in fmol.

Fig. 6.

(A) Saturation binding of ¹²⁵I-rhβ₂GPI to DS and moAb1 to β₂GPI. Saturation binding of ¹²⁵I-rhβ₂GPI (1.56-200 nM) to immobilized moAb1 (▴), DS (▾), and BSA (▪) was performed as indicated in Methods. (B) Direct Binding of ¹²⁵I-rhβ₂GPI to microtiter wells coated with PS, DS, or BSA. The amount of ¹²⁵I-rhβ₂GPI bound was determined as described in Methods.