Haemostasis 1992;22:233-235
r
. c a s ü iio
J Monteagudo J.C. Reverter A. Ordinas
Hemostatic Effect of Platelet von Willebrand Factor
Hospital Clínico, Universidad de Barcelona, Spain
Abstract
In type III von Willebrand disease (vWD) patients, the bleed ing time was only partially corrected or not modified after cryoprecipitate infusion, although the levels and the multi meric structure of plasma von Willebrand factor (vWF) were normal. However, the adhesion of normal platelets on the ves sel wall subendothelium in the presence of postinfusion pa tient plasma improved more significantly than the bleeding time. These results suggest a role of the vWF released from normal platelets which is absent in type III vWD platelets. In 5 patients transfusion of normal platelet concentrates per formed 1 h after cryoprecipitate infusion without modifica tion of the bleeding time ( > 30 min) normalized this parame ter, and platelet adhesion to the subendothelium elicited a marked improvement. These last results confirm the sugges tion that platelet vWF plays an important ‘in vivo’ role in the hemostatic process, particularly in patients suffering from severe vWD.
There are previous clinical and experimen tal suggestions that the von Willebrand factor (vWF) present in the platelet a-granules plays an important role in the hemostatic pro cess, particularly in von Willebrand disease (vWD). It was observed that type I vWD patients with normal levels of platelet vWF
have shorter bleeding times (BT) than those with low or dysfunctional platelet vWF, de spite equally low plasma vWF levels [1, 2], Pigs with severe vWD engrafted with normal bone marrow, producing vWF-containing platelets, had shorter BT than before trans plantation, despite the fact that the plasma
Dr. R. Castillo Servicio de Hemoterapia y Hemostasia Hospital Clinic Provincial Villarroel, 170 E-08036 Barcelona (Spain)
© 1992 S. Karger AG, Basel 0301-0147/92/ 0225-0233S2.75/0
Downloaded by: Karolinska Institutet, University Library 130.237.122.245 - 1/29/2019 10:12:12 AM
Key Words Type III von Willebrand disease Platelet von Willebrand factor Platelet transfusion Cryoprecipitate infusion
234
VIII activity. In 2 patients, cryoprecipitate infusion did not modify the BT ( > 30 min), whereas in the remaining 3 patients the BT was only partially corrected (from > 30 to 12, 18, and 21 min, respectively). However, the immediate platelet transfusion completely corrected the BT in 4 cases, and in 1 case it shortened the BT to 8.30 min (n = 3 to 8 min). In all patients the duration of the normaliza tion of the BT was up to 4 h after the begin ning of the platelet concentrate transfusion, and in 1 of the 4 patients it was up to 16 h. In the perfusion study, cryoprecipitate infusion only resulted in a slight increase in plate let deposition (surface coverage range 2.411.3%), whereas the platelet concentrate transfusion elicited a more marked improve ment (range 8.2-26.4%; p < 0.02 vs. postcryoprecipitate). The results obtained by the transfusion of normal platelets confirm directly the sugges tion that platelet vWF plays an important ‘in vivo’ role in supporting normal BT and platelet-vessel wall interaction. They also support the clinical indication of platelet concentrates after plasma concentrate infusion in those sit uations where this last treatment will not con trol serious hemorrhages in severe vWD. From the present data available, it cannot be settled whether the hemostatic effect of the transfused normal platelets is only due to the platelet vWF. However, there are several rea sons supporting the important role of platelet vWF in the normalization of BT and in the correction of bleeding. Platelet vWF is pre sumably the only protein brought by normal platelets that is absent in patients before transfusion. Although some degree of platelet activation has been observed during proce dure of platelet obtainment [10], in our study the BT remained normalized for more than 16 h, and the levels of platelet vWF lasted over this period after transfusion. Moreover, previous experimental data indicate that nor-
Castillo/Escolar/Monteagudo/Reverter/ Ordinas
Severe von Willebrand Disease
Downloaded by: Karolinska Institutet, University Library 130.237.122.245 - 1/29/2019 10:12:12 AM
vWF levels remained very low [3]. Moreover, it was shown that the transfusion of plasma concentrates to type III vWD patients is not able by itself to correct the BT, even though both the vWF activities and the multimeric structure of vWF were normalized [4-6], We have compared plasma levels of vWF with BT values and platelet adhesion on ves sel wall subentothelium after twelve plasma concentrate infusions, given to 4 type III vWD patients [7], Their prolonged BT were either completely or partially corrected after five infusions and did not change after admin istration of the remaining seven infusions. In contrast, the low platelet coverage of the subendothelial surface of rabbit aorta perfused with normal washed platelets and red cells resuspended in preinfusion patient plasma was completely or partially corrected in ten instances by replacing preinfusion plasma with postinfusion plasma and remained un changed in two. Postinfusion improvement in surface coverage was greater than that in BT, suggesting a role of the vWF released from normal platelets. This possibility has been further explored through other perfusion experiments [7], The subendothelial surface covered by platelets from an untreated patient with type III vWD (containing no measurable vWF) or from a type IIA vWD patient (containing dysfunc tional vWF) resuspended in normal plasma is much smaller than that covered by normal platelets resuspended in normal plasma. Sim ilar results were obtained using fibrillar colla gen as an adhesive surface [8]. Afterwards, it was attempted to prove the ‘in vivo’ hemostatic effect of normal platelets in vWD patients [9]. We have examined the effect of normal platelet concentrate transfu sion 1 h after cryoprecipitate infusion in 5 type III vWD patients. The cryoprecipitate infusion attained normal circulating levels of ristocetin cofactor, vWF antigen, and factor
mal nonactivated platelets in the presence of normal plasma adhere in vitro to collagen and to vascular subendothelium, but the adhesion of platelets from vWD patients resuspended in the same plasma is defective [7, 8], Now we are trying to specify the role of platelet vWF, particularly in the hemostatic effect of normal platelet transfusions in severe vWD. The presence of normal platelets trans fused in the aggregates on the subendothelium is evaluated by the immunogold labelling for vWF. The degree of platelet activation during platelet obtainment procedures is evaluated
by the exposure of CD62 and CD63 on the platelet surface using flow cytometry. In another aspect, we are attempting to verify which plasma vWF level is necessary after transfusion of normal platelets in type III vWD to obtain a hemostatic effect. One hour after the end of the platelet transfusion, cryoprecipitate is infused. BT, plasma and platelet vWF levels, and platelet interaction on the subendothelium are determined before and after platelet transfusion, and sequen tially during the cryoprecipitate infusion.
References 4 Mannucci PM, Moia M, Rebulla P, Altieri D, Monteagudo J, Castillo R: Correction of the bleeding time in treated patients with severe von Willebrand disease is not solely de pendent on the normal multimeric structure of plasma von Willebrand factor. Am J Hematol 1987;25:55. 5 Smiley RK, Tittley P, Rock G: Stud ies on the prolonged bleeding time in von Willebrand’s disease. Thromb Res 1989;53:147. 6 Cattaneo M, Moia M, Della Valle P. Castellana P, Mannucci PM: DDAVP shortens the prolonged bleeding times of patients with se vere von Willebrand disease treated with cryoprecipitates. Evidence for a mechanism of action independent of released von Willebrand factor. Blood 1989:74:1972.
7 Castillo R, Escolar G, Monteagudo J, Ordinas A, Garrido M, Moia M. Federici AB, Mannucci PM: Role for platelet von Willebrand factor in supporting platelet-vessel wall inter actions in von Willebrand’s disease. Am J Hematol 1989;31:153. 8 Fressinaud E, Baruch D, Rothschild C, Baumgartner JR, Meyer D: Plate let von Willebrand factor: Evidence for its involvement in platelet adhe sion to collagen. Blood 1987:70: 1214. 9 Castillo R, Monteagudo J, Escolar G, Ordinas A, Magallon M, Martin Villar J: Hemostatic effect of normal platelet transfusion in severe von Willebrand disease patients. Blood 1991;77:1901. 10 George GN, Pickett EB, Heinz R: Platelet membrane glycoprotein changes during the preparation and storage of platelet concentrates. Transfusion 1988:2:123.
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1 Mannucci PM, Lombardi R. Bader R, Vianello L, Federici AB, Solinas S, Mazzuconi MG, Mariani G: Het erogeneity of type I von Willebrand disease: Evidence for a subgroup with an abnormal von Willebrand factor. Blood 1985;66:796. 2 Gralnick HR. Rick ME, Mckeown LP, Williams SB. Parker RI, Mai sonneuve P. Jeanneau C, Sultan Y: Platelet von Willebrand factor: An important determinant of the bleed ing time in type I von Willebrand’s disease. Blood 1986;68:58. 3 Bowie EJW, Solberg LA, Fass DN, Johnson CM, Knutson GJ. Stewart ML. Zoecklein LJ: Transplantation of bone marrow into a pig with von Willebrand’s disease. J Clin Invest 1986;78:26.
r
. c a s ü iio
J Monteagudo J.C. Reverter A. Ordinas
Hemostatic Effect of Platelet von Willebrand Factor
Hospital Clínico, Universidad de Barcelona, Spain
Abstract
In type III von Willebrand disease (vWD) patients, the bleed ing time was only partially corrected or not modified after cryoprecipitate infusion, although the levels and the multi meric structure of plasma von Willebrand factor (vWF) were normal. However, the adhesion of normal platelets on the ves sel wall subendothelium in the presence of postinfusion pa tient plasma improved more significantly than the bleeding time. These results suggest a role of the vWF released from normal platelets which is absent in type III vWD platelets. In 5 patients transfusion of normal platelet concentrates per formed 1 h after cryoprecipitate infusion without modifica tion of the bleeding time ( > 30 min) normalized this parame ter, and platelet adhesion to the subendothelium elicited a marked improvement. These last results confirm the sugges tion that platelet vWF plays an important ‘in vivo’ role in the hemostatic process, particularly in patients suffering from severe vWD.
There are previous clinical and experimen tal suggestions that the von Willebrand factor (vWF) present in the platelet a-granules plays an important role in the hemostatic pro cess, particularly in von Willebrand disease (vWD). It was observed that type I vWD patients with normal levels of platelet vWF
have shorter bleeding times (BT) than those with low or dysfunctional platelet vWF, de spite equally low plasma vWF levels [1, 2], Pigs with severe vWD engrafted with normal bone marrow, producing vWF-containing platelets, had shorter BT than before trans plantation, despite the fact that the plasma
Dr. R. Castillo Servicio de Hemoterapia y Hemostasia Hospital Clinic Provincial Villarroel, 170 E-08036 Barcelona (Spain)
© 1992 S. Karger AG, Basel 0301-0147/92/ 0225-0233S2.75/0
Downloaded by: Karolinska Institutet, University Library 130.237.122.245 - 1/29/2019 10:12:12 AM
Key Words Type III von Willebrand disease Platelet von Willebrand factor Platelet transfusion Cryoprecipitate infusion
234
VIII activity. In 2 patients, cryoprecipitate infusion did not modify the BT ( > 30 min), whereas in the remaining 3 patients the BT was only partially corrected (from > 30 to 12, 18, and 21 min, respectively). However, the immediate platelet transfusion completely corrected the BT in 4 cases, and in 1 case it shortened the BT to 8.30 min (n = 3 to 8 min). In all patients the duration of the normaliza tion of the BT was up to 4 h after the begin ning of the platelet concentrate transfusion, and in 1 of the 4 patients it was up to 16 h. In the perfusion study, cryoprecipitate infusion only resulted in a slight increase in plate let deposition (surface coverage range 2.411.3%), whereas the platelet concentrate transfusion elicited a more marked improve ment (range 8.2-26.4%; p < 0.02 vs. postcryoprecipitate). The results obtained by the transfusion of normal platelets confirm directly the sugges tion that platelet vWF plays an important ‘in vivo’ role in supporting normal BT and platelet-vessel wall interaction. They also support the clinical indication of platelet concentrates after plasma concentrate infusion in those sit uations where this last treatment will not con trol serious hemorrhages in severe vWD. From the present data available, it cannot be settled whether the hemostatic effect of the transfused normal platelets is only due to the platelet vWF. However, there are several rea sons supporting the important role of platelet vWF in the normalization of BT and in the correction of bleeding. Platelet vWF is pre sumably the only protein brought by normal platelets that is absent in patients before transfusion. Although some degree of platelet activation has been observed during proce dure of platelet obtainment [10], in our study the BT remained normalized for more than 16 h, and the levels of platelet vWF lasted over this period after transfusion. Moreover, previous experimental data indicate that nor-
Castillo/Escolar/Monteagudo/Reverter/ Ordinas
Severe von Willebrand Disease
Downloaded by: Karolinska Institutet, University Library 130.237.122.245 - 1/29/2019 10:12:12 AM
vWF levels remained very low [3]. Moreover, it was shown that the transfusion of plasma concentrates to type III vWD patients is not able by itself to correct the BT, even though both the vWF activities and the multimeric structure of vWF were normalized [4-6], We have compared plasma levels of vWF with BT values and platelet adhesion on ves sel wall subentothelium after twelve plasma concentrate infusions, given to 4 type III vWD patients [7], Their prolonged BT were either completely or partially corrected after five infusions and did not change after admin istration of the remaining seven infusions. In contrast, the low platelet coverage of the subendothelial surface of rabbit aorta perfused with normal washed platelets and red cells resuspended in preinfusion patient plasma was completely or partially corrected in ten instances by replacing preinfusion plasma with postinfusion plasma and remained un changed in two. Postinfusion improvement in surface coverage was greater than that in BT, suggesting a role of the vWF released from normal platelets. This possibility has been further explored through other perfusion experiments [7], The subendothelial surface covered by platelets from an untreated patient with type III vWD (containing no measurable vWF) or from a type IIA vWD patient (containing dysfunc tional vWF) resuspended in normal plasma is much smaller than that covered by normal platelets resuspended in normal plasma. Sim ilar results were obtained using fibrillar colla gen as an adhesive surface [8]. Afterwards, it was attempted to prove the ‘in vivo’ hemostatic effect of normal platelets in vWD patients [9]. We have examined the effect of normal platelet concentrate transfu sion 1 h after cryoprecipitate infusion in 5 type III vWD patients. The cryoprecipitate infusion attained normal circulating levels of ristocetin cofactor, vWF antigen, and factor
mal nonactivated platelets in the presence of normal plasma adhere in vitro to collagen and to vascular subendothelium, but the adhesion of platelets from vWD patients resuspended in the same plasma is defective [7, 8], Now we are trying to specify the role of platelet vWF, particularly in the hemostatic effect of normal platelet transfusions in severe vWD. The presence of normal platelets trans fused in the aggregates on the subendothelium is evaluated by the immunogold labelling for vWF. The degree of platelet activation during platelet obtainment procedures is evaluated
by the exposure of CD62 and CD63 on the platelet surface using flow cytometry. In another aspect, we are attempting to verify which plasma vWF level is necessary after transfusion of normal platelets in type III vWD to obtain a hemostatic effect. One hour after the end of the platelet transfusion, cryoprecipitate is infused. BT, plasma and platelet vWF levels, and platelet interaction on the subendothelium are determined before and after platelet transfusion, and sequen tially during the cryoprecipitate infusion.
References 4 Mannucci PM, Moia M, Rebulla P, Altieri D, Monteagudo J, Castillo R: Correction of the bleeding time in treated patients with severe von Willebrand disease is not solely de pendent on the normal multimeric structure of plasma von Willebrand factor. Am J Hematol 1987;25:55. 5 Smiley RK, Tittley P, Rock G: Stud ies on the prolonged bleeding time in von Willebrand’s disease. Thromb Res 1989;53:147. 6 Cattaneo M, Moia M, Della Valle P. Castellana P, Mannucci PM: DDAVP shortens the prolonged bleeding times of patients with se vere von Willebrand disease treated with cryoprecipitates. Evidence for a mechanism of action independent of released von Willebrand factor. Blood 1989:74:1972.
7 Castillo R, Escolar G, Monteagudo J, Ordinas A, Garrido M, Moia M. Federici AB, Mannucci PM: Role for platelet von Willebrand factor in supporting platelet-vessel wall inter actions in von Willebrand’s disease. Am J Hematol 1989;31:153. 8 Fressinaud E, Baruch D, Rothschild C, Baumgartner JR, Meyer D: Plate let von Willebrand factor: Evidence for its involvement in platelet adhe sion to collagen. Blood 1987:70: 1214. 9 Castillo R, Monteagudo J, Escolar G, Ordinas A, Magallon M, Martin Villar J: Hemostatic effect of normal platelet transfusion in severe von Willebrand disease patients. Blood 1991;77:1901. 10 George GN, Pickett EB, Heinz R: Platelet membrane glycoprotein changes during the preparation and storage of platelet concentrates. Transfusion 1988:2:123.
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1 Mannucci PM, Lombardi R. Bader R, Vianello L, Federici AB, Solinas S, Mazzuconi MG, Mariani G: Het erogeneity of type I von Willebrand disease: Evidence for a subgroup with an abnormal von Willebrand factor. Blood 1985;66:796. 2 Gralnick HR. Rick ME, Mckeown LP, Williams SB. Parker RI, Mai sonneuve P. Jeanneau C, Sultan Y: Platelet von Willebrand factor: An important determinant of the bleed ing time in type I von Willebrand’s disease. Blood 1986;68:58. 3 Bowie EJW, Solberg LA, Fass DN, Johnson CM, Knutson GJ. Stewart ML. Zoecklein LJ: Transplantation of bone marrow into a pig with von Willebrand’s disease. J Clin Invest 1986;78:26.
