THROMBOSIS RESEARCH 67; 23-30,1992 0049-3848/92 $5.00 + .OOPrinted in the USA. Copyright (c) 1992 Pergamon Press Ltd. All rights reserved.
COMBINED INHERITED PROTEIN S AND HEPARIN CO-FACTOR II DEFICIENCY IN A PATIENT WITH UPPER LIMB THROMBOSIS: A FAMILY STUDY. Simioni P., Zanardi S., Prandoni P.* and Girolami A. Institute of Medical Semeiotics, 2nd Chair of Medicine and * Second Department of Internal Medicine University of Padua Medical School, Padua, Italy. (Received 11.l .1992; accepted in revised form 13.4.1992 by Editor M.F. Scully) (Received by Executive Editorial Office 1.6.1992)
ABSTRACT
A 42-year-old Italian woman presenting with spontaneous deep vein thrombosis of the right arm, was found to have inherited a deficiency of both protein S (PS) and heparin co-factor II (HC II). The two defects seemed to segregate independently, since her son exhibited only a HC II deficiency while one of her sisters manifested only the PS defect. All affected patients appeared heterozygous for one or other or both deficiency states. The proposita and her sister exhibited a congenital PS deficiency consisting of normal or near normal levels of total PS antigen and C4b-binding protein (CQb-BP) but a moderate reduction both of free PS antigen and of PS functional activity. In addition, the proposita and her son had half normal levels of HC II antigen and activity. Except for the proposita,all were asymptomatic. Inherited deficiencies either of PS or of HC II have been associated with thrombotic manifestations. Since the proposita had an inherited combined defect of the two proteins,severe thrombotic events might be expected. However, this was not found to be the case. The role of HC II deficiency in the pathogenesis of thrombosis whether alone or combined remains to be fully investigated.
Key words: combined hereditary defect, thrombophilia, protein S deficiency, heparin cofactor II deficiency, deep venous thrombosis (DVT), upper limbs thrombosis
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INTRODUCTION Combined congenital defects of coagulation inhibitor@ (antithrombin III, protein C, protein S, heparin co-factor II) has been reported only in a few families(1,2,3,4). This is probely due to the fact that the coincidental inheritance of more than one deficiency state is a highly unusual occurrence. Since single congenital defects have been associated with an increased incidence of thrombotic manifestations (5-l()), combination of two or more defects could be maintained to give more severe thrombophilic state. This was not found to be always the case, especially when the affected patients were young (4). Here we describe the case of patient with a congenital combined defect of HC II and protein S who remained asymptomatic until 42 of DVT of right arm. The age when she developed a spontaneous inheritance of the defect by the laboratory was confirmed investigation of some family members. CASE REPORT A 42-year-old Italian woman presenting with spontaneous deep vein thrombosis of the right arm, was admitted to our internal medicine department last year. She complained of pain and swelling to the right arm. A diagnosis of right axillary-vein thrombosis was confirmed by echo-doppler and venography. Treatment with standard heparin followed by three months oral anticoagulation were effective and the patient had a complete recovery. Laboratory study was performed in the proposita six months after dismissal of oral anticoagulant treatment. In the follow-up of the patient, chest CTscan and right arm venography were performed and no anatomical cause of upper limb thrombosis was found. The fbmily members available for study were also investigated. They all ware asymptomatic. MATERIALS
AND METHODS
Routine coagulation tests employed have been reported previously (8,9,10). Normal ranges were considered as the mean+/-2SD of assays carried out in 30 normal subjects of both sexes aged 18-50. Briefly, AT III activity measurement was performed in plasma using two chromogenic substrates (9). AT III antigen was evaluated by rocket electrophoresis according to the method of Laurel1 (ll), using anti-AT111 antiserum supplied by Behringwerke, Marburq, Germany, at a concentration of 2.5% . The coagulometric assay for Protein C activity was performed using the "Protein C reagent" provided by Behringwerke (Marburg, Germany), based on protein C activation by Protac. Protein C antigen level was evaluated according to the method of Laurel1 (11) using agarose 1% in Na Barbital buffer at pH 8.8 and containing anti-protein C antiserum at a concentration of 1.2% supplied by Stago,Asnieres,France.
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Protein S assay. -Total protein S antigen was determined by Laurel1 rocket electrophoresis (10,ll) carried out using agarose 1% in Na Barbital buffer pH 8.8, containing protein-S antiserum supplied by Stago at a concentration of 0.8% : plasma samples of 5 ul were used and electrophoresed at 1 mA/cm for 20 hrs with the cooling bath set at 18" C. Normal ranges were 99.5 +/-29.8% (mean+/-2SD). -Free protein S antigen was quantified after precipitation of the C4b-BP-protein S complex by Polyethylene Glycol (PEG) 6000 (10). The supernatant was analyzed by Laurel1 rocket electrophoresis (11) under the same conditions as described for total protein S antigen. Normal ranges were 102+/-25.8% (mean+/-2SD). -Crossed immunoelectrophoresis (CIEP) of total protein S and free protein S (supernatant of PEG-treated plasma) was performed using agarose 1% in Na Barbital buffer pH 8.5 . The first migration was carried out at 14" C, at 1 mA/cm for 4 hours. The second migration was obtained in agarose 1% containing 2% PEG 6000 and protein-S antiserum (supplied by Stago) at a concentration of 1.2% , applying lmA/cm for 6 hours at room temperature. -Protein S activity was assayed using the Staclot Protein S, supplied by Stago which is based upon protein S cofactor activity in the anticoagulant action of activated protein C (APC) in a system enriched with factor Va as physiological substrate of APC (12). Normal ranges were lOl+/-29.9% (mean+/-2SD). -C4b-binding protein (C4b-BP) was evaluated by Laurel1 (11) using agarose 1% in Na Barbital buffer pH 8.8 containing anti-C4b-BP antiserum,supplied by Stago, at a concentration of 1% .Normal ranges were 98+/-28.4% (mean+/-2SD). Heparin Cofactor II assay. -Heparin Cofactor II (HC II) antigen was evaluated according to the method of Laurel1 (11) under the conditions reported previously (8). Normal ranges were 98+/-31.8% (mean+/-2SD). -Heparin Cofactor II activity was measured using the Stachrom HC II provided by Stago (8). The method is mainly based on the inhibition of thrombin by HC II after dilution of plasma samples in a The remaining activity of thrombin in dermatan sulfate solution. plasma is then detected by evaluating its amidolytic effect on a chromogenic substrate. Normal ranges were 102+/-30.9% (mean+/-2SD). the proposita's plasma was -Crossed immunoelectrophoresis of performed as reported in a previous paper (8).
RESULTS The proposita and several of family members (see family pedigree) were investigated (Fig-l) but not all relatives were available for study. levels of the family members Table 1 shows PS, HC II and C4b-BP investigated. The proposita (11-2) had a slight reduction of total PS antigen levels of free PS antigen (49%) and (60%) and half the normal the normal range activity (52%) while C4b-BP level was within (70%). The proposita's sister (11-l) had a normal total PS antigen (60%) and PS value (90%) and slightly reduced free PS antigen activity (63%), with a normal level of C4b-BP (82%). These findings
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were consistent with PS deficiency in the proposita and in her sister. The two sons of the proposita were found to possess normal levels of PS and C4b-BP. CIEP of the proposita's plasma showed a slightly reduced peak of PS-C4b-BP complex and moderately reduced free PS antigen peak (more with pooled anodal) as compared normal After PEGplasma. precipitation of PS-C4b-BP complex, CIEP of supernatant again showed reduced level of free PS in the proposita when compared with pooled normal plasma. A similar CIEP pattern was found in the proposita's sister (II-l). In this patient, however, the PS-C4b-BP complex peak was normal. Both HC II antigen and activity were found to be reduced in the proposita (II-2)(47% and 60%, respectively) and in her son (111-2) (48% and 58%, respectively) (Table 1). These findings were consistent with an inherited heterozygous HC II state. The other patients investigated had normal levels of HC II antigen and activity. CIEP of HC II of showed the plasma normal proposita's electrophoretic mobility although the peak appeared to be smaller when compared with pooled normal plasma. All the other routine coagulation tests performed in the proposita and in his family members were within normal ranges.
TABLE 1 Main laboratory
features of the family members
investigated.
PROTEIN S
legend: AG=antigen; ACT=activity; HC II=Heparin Cofactor C4b-BP = C4b-binding protein
II;
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CO-FACTOR II DEFICIENCY
HC IIdeficency
? ? ?S Protein
FIG.
deficency Defect
1
Family pedigree. The proposita is indicated by the arrow. Patients III-l, 111-2, 11-l and II-2 were 17, 21, 60 and 42 of age, respectively.
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DISCUSSION Hereditary deficiency of blood coagulation inhibitors,i.e., AT III, protein C, protein S is associated with an increased incidence of thrombotic manifestations in affected patients (5,13,14). The postulated association between hereditary HC II defects and venous thrombosis is still under investigation (15), although several possible cases have been reported (6,7,8). Combined defects of coagulation factors in which at least one of the deficient proteins was a clotting inhibitor have already been reported (16-21). Patients with congenital combined defects of Von Willebrand factor and antithrombin III (16) have been found to be asymptomatic possibly by virtue of a protective effect of the former against the latter. On the other hand, progressive pulmonary hypertension and hemoptysis have been described in a patient with combined protein C and factor VII defects congenital (17). inherited dysfibrinogenaemia protein C Association of and deficiency (18), homozygous protein C deficiency and heterozygous dysplasminogenemia (19), antithrombin III deficiency and factor XII deficiency (20), have also been reported in thrombophilic patients. Similarly combined deficiencies of two coagulation inhibitors have also been found namely, combined protein C and AT III deficiency or abnormality (1,2,3) and combined protein S and AT III deficiency (1). Recently, combined defects of protein S and AT III, HC II and protein C or protein S, protein C and protein S have all been reported in a large kindred (4). In the same family two patients had triple defects, namely combined AT III - HC II - Protein C deficiency and combined AT III - HC II -Protein S deficiency (4). In the case here described, an association of inherited protein S and HC II deficiencies was found in a female who experienced a DVT of the right arm at the age of 40. No other risk factors for thrombosis such as oral contraceptive treatment, smoking, surgery or trauma were present in the proposita. The patient had half the normal level of free protein S, with normal levels of total protein S and C4B-binding protein. The same defect was found in her sister. Crossed immunoelectrophoresis confirmed the abnormal distribution of protein S in the proposita and in her sister. In addition, the proposita had concurrently reduced levels of HC II antigen and activity consistent with a type I heterozygous defect. The same defect was detected in one of her sons. None of the patients had vitamin-K deficiency or other acquired conditions such as liver disease or disseminated intravascular coagulation (DIC) which could explain the laboratory findings. The data suggest that the two hereditary defects segregated independently. Symptoms were present only in the proposita while patients with a single defect were asymptomatic. The proposita experienced her first and only thrombotic episode at the age of 40. This seems to be in contrast to other reports on combined defects of coagulation inhibitors, where the affected patients experienced severe venous thrombotic episodes (1,2). However, the role of HC II deficiency in the pathogenesis of thrombosis whether alone or combined remains to be fully investigated. In fact the frequency of HC II deficiency in patient with DVT is not significantly different from the frequency in healthy control populations (15). Perhaps this study gives some way to suggest that HC II deficiency may facilitate the expression of a thrombotic state in a patient with a second,different deficiency state.
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29
REFERENCES 1.
BAUER, K.A., BROEKMANS, A.W., BERTINA, R.M., CONARP, J., HORELLOU, M-H., SAMAMA, M.M., ROSENBERG, R.D. He-static enzyme generation in the blood of patients with hereditary protein C deficiency. Blood, 71, 1418-1426, 1988.
2. SAS, G., PAL, A. Severe familial thrombophilia caused by the combination of protein C deficiency and minor qualitative disturbance of antithrombin III production. Thromb. Haemorrh. Disorders, 1, 29-32, 1990. 3. WOLF, M., BOYER-NEUMANN, C., MOLHO-SABATIER, P., NEUMANN, MEYER, D., LARRIEU, M.J. Familial variant of antithrombin (AT III Bligny,47 Arg to His) associated with protein C deficiency. Thromb. Haemost., 63, 215-219, 1990.
C., III
4. JOBIN, F., VU, L., LESSARD, M. Two cases of inherited triple deficiency in a large kindred with thrombotic diathesis and deficiencies of antithrombin III, Heparin Cofactor II, protein C and protein S. Thromb. Haemost., 66, 295-299, 1991. 5. COMP, P-C., NIXON, R.R., COOPER, M.R., ESMON, C.T. Familial protein S deficiency is associated with recurrent thrombosis. J. Clin. Invest., 74, 2082-2088, 1984. 6. SIE, P., DUPOUY, D., PICHON, J., BONEU, P. Constitutional heparin co-factor II deficiency associated to recurrent venous thrombosis. Lancet, ii, 414-416, 1985. 7. TRAN, T-H., MARBET, G-A., DUCKERT, F. Association of hereditary heparin co-factor II deficiency with thrombosis. Lancet, . . 11, 413-414, 1985. P., LAZZARO, A.R., COSER, E., SALMISTRARO, G., 8. SIMIONI, GIRdLAMI, A. Hereditary heparin cofactor II deficiency and thrombosis: report of six patients belonging to two separate kindreds. Blood Coaqulation and Fibrinolysis, 1, 351-356, 1990. M.G., VICARIOTO, M.A., MARAFIOTI, A., CAPPELLATO, 9. GIROLAMI, F ., TRAVERSO, R., VERGOLANI, A., BOERI, G. Antithrombin III a report of 14 cases belonging to three different deficiency: Folia Haematol., 112, 594-606,1985. kindreds. lO.GIROLAMI, A., SIMIONI, P., LAZZARO, A.R., CORDIANO, I. Severe arterial cerebral thrombosis in a patient with protein S deficiency (moderately reduced total and markedly reduced free protein S):a family study. Thromb. Haemost., 61, 144-147,1989. ll.LAURELL, phoresis 1968.
C.B. Quantitative estimation of proteins of human serum proteins. Anal. Biochem.,
by electro35, 403-413,
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BOYER-NEUMANN, C., MARTINOLI, J.L., LEROY-MATHERON, 12.WOLF, M., J., MEYER, D., LARRIEU, M.J. A new functional C ., AMIRAL, assay for human protein S activity using activated factor V as substrate. Thromb. Haemost., 62, 1144-1145, 1989. 13.EGEBERG, 0. Inherited antithrombin III deficiency causing Thromb. Diath. Hemorrh., 13, 516-530, 1965. thrombophilia. 14.GRIFFIN, WIEDEMAN, disease.
J.H., EVATT, B., ZIMMERMANN, T.S., KLEISS, A.J., C. Deficiency of protein C in congenital thrombotic J. Clin. Invest., 68, 1370-1373, 1981.
L., MULLER, 15.BERTINA, R.M., VAN DER LINDEN, I.K., ENGESSER, E.J.P. Hereditary heparin cofactor II H.P., BROMMER, deficiency and the risk of development of thrombosis. Thromb.Haemost., 57, 196-200, 1987. 16.GIROLAM1, A., CAPPELLATO, M.G., VICARIOTO, M.A., CASONATO, S., F. Associated von Willebrand disease as a possible MARAFIOTI, cause of lack of thrombosis in an AT III abnormality (AT III Trento). Blut, 52, 29-33, 1986. 17.TAKEUCH1, Y., SAITO, Y., IKENOUCHI, H., SUGIMOTO, T., SUGOH, T ., SAKATA, Y., MATSUDA, M., TAKAMIYA, 0. Combined factor VII and protein C deficiency with peripheral pulmonary artery stenosis accompanied by progressive pulmonary hypertension and hemoptysis. Thromb. Res., 51, 117-126, 1988. 18.GRANDILLE, S., PRIOLLET, P., CAPRON, L., RONCATO, M., of inherited FIESSINGER, J.N., AIACH, M. Association dysfibrinogenaemia and protein C deficiency in two unrelated families. Br. J. Haemat., 68, 329-337, 1988. lg.MANABE, S., MATSUDA, M. Homozygous protein C deficiency combined with heterozygous dysplasminogenemia found in a 21 -year-old thrombophilic male. Thromb. Res., 39, 333-341, 1985. 2O.MAURIN, N., SCHUNKERT, H., combination of antithrombin Thromb. Haemost., 60, 127,
SIEBERTH, H.G. Thrombophilia with III and factor XII deficiency. 1988.
a
21.BRENNER, B., TAVORI, S., ZIVELIN, A., KELLER, C.B., SUTTIE, I., SELIGSOHN, J.W., TATARSKY, U. Hereditary deficiency of all vitamin K-dependent procoagulants and anticoagulants. Br. J. Haemat., 75, 537-542, 1990.
COMBINED INHERITED PROTEIN S AND HEPARIN CO-FACTOR II DEFICIENCY IN A PATIENT WITH UPPER LIMB THROMBOSIS: A FAMILY STUDY. Simioni P., Zanardi S., Prandoni P.* and Girolami A. Institute of Medical Semeiotics, 2nd Chair of Medicine and * Second Department of Internal Medicine University of Padua Medical School, Padua, Italy. (Received 11.l .1992; accepted in revised form 13.4.1992 by Editor M.F. Scully) (Received by Executive Editorial Office 1.6.1992)
ABSTRACT
A 42-year-old Italian woman presenting with spontaneous deep vein thrombosis of the right arm, was found to have inherited a deficiency of both protein S (PS) and heparin co-factor II (HC II). The two defects seemed to segregate independently, since her son exhibited only a HC II deficiency while one of her sisters manifested only the PS defect. All affected patients appeared heterozygous for one or other or both deficiency states. The proposita and her sister exhibited a congenital PS deficiency consisting of normal or near normal levels of total PS antigen and C4b-binding protein (CQb-BP) but a moderate reduction both of free PS antigen and of PS functional activity. In addition, the proposita and her son had half normal levels of HC II antigen and activity. Except for the proposita,all were asymptomatic. Inherited deficiencies either of PS or of HC II have been associated with thrombotic manifestations. Since the proposita had an inherited combined defect of the two proteins,severe thrombotic events might be expected. However, this was not found to be the case. The role of HC II deficiency in the pathogenesis of thrombosis whether alone or combined remains to be fully investigated.
Key words: combined hereditary defect, thrombophilia, protein S deficiency, heparin cofactor II deficiency, deep venous thrombosis (DVT), upper limbs thrombosis
23
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INTRODUCTION Combined congenital defects of coagulation inhibitor@ (antithrombin III, protein C, protein S, heparin co-factor II) has been reported only in a few families(1,2,3,4). This is probely due to the fact that the coincidental inheritance of more than one deficiency state is a highly unusual occurrence. Since single congenital defects have been associated with an increased incidence of thrombotic manifestations (5-l()), combination of two or more defects could be maintained to give more severe thrombophilic state. This was not found to be always the case, especially when the affected patients were young (4). Here we describe the case of patient with a congenital combined defect of HC II and protein S who remained asymptomatic until 42 of DVT of right arm. The age when she developed a spontaneous inheritance of the defect by the laboratory was confirmed investigation of some family members. CASE REPORT A 42-year-old Italian woman presenting with spontaneous deep vein thrombosis of the right arm, was admitted to our internal medicine department last year. She complained of pain and swelling to the right arm. A diagnosis of right axillary-vein thrombosis was confirmed by echo-doppler and venography. Treatment with standard heparin followed by three months oral anticoagulation were effective and the patient had a complete recovery. Laboratory study was performed in the proposita six months after dismissal of oral anticoagulant treatment. In the follow-up of the patient, chest CTscan and right arm venography were performed and no anatomical cause of upper limb thrombosis was found. The fbmily members available for study were also investigated. They all ware asymptomatic. MATERIALS
AND METHODS
Routine coagulation tests employed have been reported previously (8,9,10). Normal ranges were considered as the mean+/-2SD of assays carried out in 30 normal subjects of both sexes aged 18-50. Briefly, AT III activity measurement was performed in plasma using two chromogenic substrates (9). AT III antigen was evaluated by rocket electrophoresis according to the method of Laurel1 (ll), using anti-AT111 antiserum supplied by Behringwerke, Marburq, Germany, at a concentration of 2.5% . The coagulometric assay for Protein C activity was performed using the "Protein C reagent" provided by Behringwerke (Marburg, Germany), based on protein C activation by Protac. Protein C antigen level was evaluated according to the method of Laurel1 (11) using agarose 1% in Na Barbital buffer at pH 8.8 and containing anti-protein C antiserum at a concentration of 1.2% supplied by Stago,Asnieres,France.
Vol. 67, No. 1
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25
Protein S assay. -Total protein S antigen was determined by Laurel1 rocket electrophoresis (10,ll) carried out using agarose 1% in Na Barbital buffer pH 8.8, containing protein-S antiserum supplied by Stago at a concentration of 0.8% : plasma samples of 5 ul were used and electrophoresed at 1 mA/cm for 20 hrs with the cooling bath set at 18" C. Normal ranges were 99.5 +/-29.8% (mean+/-2SD). -Free protein S antigen was quantified after precipitation of the C4b-BP-protein S complex by Polyethylene Glycol (PEG) 6000 (10). The supernatant was analyzed by Laurel1 rocket electrophoresis (11) under the same conditions as described for total protein S antigen. Normal ranges were 102+/-25.8% (mean+/-2SD). -Crossed immunoelectrophoresis (CIEP) of total protein S and free protein S (supernatant of PEG-treated plasma) was performed using agarose 1% in Na Barbital buffer pH 8.5 . The first migration was carried out at 14" C, at 1 mA/cm for 4 hours. The second migration was obtained in agarose 1% containing 2% PEG 6000 and protein-S antiserum (supplied by Stago) at a concentration of 1.2% , applying lmA/cm for 6 hours at room temperature. -Protein S activity was assayed using the Staclot Protein S, supplied by Stago which is based upon protein S cofactor activity in the anticoagulant action of activated protein C (APC) in a system enriched with factor Va as physiological substrate of APC (12). Normal ranges were lOl+/-29.9% (mean+/-2SD). -C4b-binding protein (C4b-BP) was evaluated by Laurel1 (11) using agarose 1% in Na Barbital buffer pH 8.8 containing anti-C4b-BP antiserum,supplied by Stago, at a concentration of 1% .Normal ranges were 98+/-28.4% (mean+/-2SD). Heparin Cofactor II assay. -Heparin Cofactor II (HC II) antigen was evaluated according to the method of Laurel1 (11) under the conditions reported previously (8). Normal ranges were 98+/-31.8% (mean+/-2SD). -Heparin Cofactor II activity was measured using the Stachrom HC II provided by Stago (8). The method is mainly based on the inhibition of thrombin by HC II after dilution of plasma samples in a The remaining activity of thrombin in dermatan sulfate solution. plasma is then detected by evaluating its amidolytic effect on a chromogenic substrate. Normal ranges were 102+/-30.9% (mean+/-2SD). the proposita's plasma was -Crossed immunoelectrophoresis of performed as reported in a previous paper (8).
RESULTS The proposita and several of family members (see family pedigree) were investigated (Fig-l) but not all relatives were available for study. levels of the family members Table 1 shows PS, HC II and C4b-BP investigated. The proposita (11-2) had a slight reduction of total PS antigen levels of free PS antigen (49%) and (60%) and half the normal the normal range activity (52%) while C4b-BP level was within (70%). The proposita's sister (11-l) had a normal total PS antigen (60%) and PS value (90%) and slightly reduced free PS antigen activity (63%), with a normal level of C4b-BP (82%). These findings
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were consistent with PS deficiency in the proposita and in her sister. The two sons of the proposita were found to possess normal levels of PS and C4b-BP. CIEP of the proposita's plasma showed a slightly reduced peak of PS-C4b-BP complex and moderately reduced free PS antigen peak (more with pooled anodal) as compared normal After PEGplasma. precipitation of PS-C4b-BP complex, CIEP of supernatant again showed reduced level of free PS in the proposita when compared with pooled normal plasma. A similar CIEP pattern was found in the proposita's sister (II-l). In this patient, however, the PS-C4b-BP complex peak was normal. Both HC II antigen and activity were found to be reduced in the proposita (II-2)(47% and 60%, respectively) and in her son (111-2) (48% and 58%, respectively) (Table 1). These findings were consistent with an inherited heterozygous HC II state. The other patients investigated had normal levels of HC II antigen and activity. CIEP of HC II of showed the plasma normal proposita's electrophoretic mobility although the peak appeared to be smaller when compared with pooled normal plasma. All the other routine coagulation tests performed in the proposita and in his family members were within normal ranges.
TABLE 1 Main laboratory
features of the family members
investigated.
PROTEIN S
legend: AG=antigen; ACT=activity; HC II=Heparin Cofactor C4b-BP = C4b-binding protein
II;
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CO-FACTOR II DEFICIENCY
HC IIdeficency
? ? ?S Protein
FIG.
deficency Defect
1
Family pedigree. The proposita is indicated by the arrow. Patients III-l, 111-2, 11-l and II-2 were 17, 21, 60 and 42 of age, respectively.
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DISCUSSION Hereditary deficiency of blood coagulation inhibitors,i.e., AT III, protein C, protein S is associated with an increased incidence of thrombotic manifestations in affected patients (5,13,14). The postulated association between hereditary HC II defects and venous thrombosis is still under investigation (15), although several possible cases have been reported (6,7,8). Combined defects of coagulation factors in which at least one of the deficient proteins was a clotting inhibitor have already been reported (16-21). Patients with congenital combined defects of Von Willebrand factor and antithrombin III (16) have been found to be asymptomatic possibly by virtue of a protective effect of the former against the latter. On the other hand, progressive pulmonary hypertension and hemoptysis have been described in a patient with combined protein C and factor VII defects congenital (17). inherited dysfibrinogenaemia protein C Association of and deficiency (18), homozygous protein C deficiency and heterozygous dysplasminogenemia (19), antithrombin III deficiency and factor XII deficiency (20), have also been reported in thrombophilic patients. Similarly combined deficiencies of two coagulation inhibitors have also been found namely, combined protein C and AT III deficiency or abnormality (1,2,3) and combined protein S and AT III deficiency (1). Recently, combined defects of protein S and AT III, HC II and protein C or protein S, protein C and protein S have all been reported in a large kindred (4). In the same family two patients had triple defects, namely combined AT III - HC II - Protein C deficiency and combined AT III - HC II -Protein S deficiency (4). In the case here described, an association of inherited protein S and HC II deficiencies was found in a female who experienced a DVT of the right arm at the age of 40. No other risk factors for thrombosis such as oral contraceptive treatment, smoking, surgery or trauma were present in the proposita. The patient had half the normal level of free protein S, with normal levels of total protein S and C4B-binding protein. The same defect was found in her sister. Crossed immunoelectrophoresis confirmed the abnormal distribution of protein S in the proposita and in her sister. In addition, the proposita had concurrently reduced levels of HC II antigen and activity consistent with a type I heterozygous defect. The same defect was detected in one of her sons. None of the patients had vitamin-K deficiency or other acquired conditions such as liver disease or disseminated intravascular coagulation (DIC) which could explain the laboratory findings. The data suggest that the two hereditary defects segregated independently. Symptoms were present only in the proposita while patients with a single defect were asymptomatic. The proposita experienced her first and only thrombotic episode at the age of 40. This seems to be in contrast to other reports on combined defects of coagulation inhibitors, where the affected patients experienced severe venous thrombotic episodes (1,2). However, the role of HC II deficiency in the pathogenesis of thrombosis whether alone or combined remains to be fully investigated. In fact the frequency of HC II deficiency in patient with DVT is not significantly different from the frequency in healthy control populations (15). Perhaps this study gives some way to suggest that HC II deficiency may facilitate the expression of a thrombotic state in a patient with a second,different deficiency state.
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29
REFERENCES 1.
BAUER, K.A., BROEKMANS, A.W., BERTINA, R.M., CONARP, J., HORELLOU, M-H., SAMAMA, M.M., ROSENBERG, R.D. He-static enzyme generation in the blood of patients with hereditary protein C deficiency. Blood, 71, 1418-1426, 1988.
2. SAS, G., PAL, A. Severe familial thrombophilia caused by the combination of protein C deficiency and minor qualitative disturbance of antithrombin III production. Thromb. Haemorrh. Disorders, 1, 29-32, 1990. 3. WOLF, M., BOYER-NEUMANN, C., MOLHO-SABATIER, P., NEUMANN, MEYER, D., LARRIEU, M.J. Familial variant of antithrombin (AT III Bligny,47 Arg to His) associated with protein C deficiency. Thromb. Haemost., 63, 215-219, 1990.
C., III
4. JOBIN, F., VU, L., LESSARD, M. Two cases of inherited triple deficiency in a large kindred with thrombotic diathesis and deficiencies of antithrombin III, Heparin Cofactor II, protein C and protein S. Thromb. Haemost., 66, 295-299, 1991. 5. COMP, P-C., NIXON, R.R., COOPER, M.R., ESMON, C.T. Familial protein S deficiency is associated with recurrent thrombosis. J. Clin. Invest., 74, 2082-2088, 1984. 6. SIE, P., DUPOUY, D., PICHON, J., BONEU, P. Constitutional heparin co-factor II deficiency associated to recurrent venous thrombosis. Lancet, ii, 414-416, 1985. 7. TRAN, T-H., MARBET, G-A., DUCKERT, F. Association of hereditary heparin co-factor II deficiency with thrombosis. Lancet, . . 11, 413-414, 1985. P., LAZZARO, A.R., COSER, E., SALMISTRARO, G., 8. SIMIONI, GIRdLAMI, A. Hereditary heparin cofactor II deficiency and thrombosis: report of six patients belonging to two separate kindreds. Blood Coaqulation and Fibrinolysis, 1, 351-356, 1990. M.G., VICARIOTO, M.A., MARAFIOTI, A., CAPPELLATO, 9. GIROLAMI, F ., TRAVERSO, R., VERGOLANI, A., BOERI, G. Antithrombin III a report of 14 cases belonging to three different deficiency: Folia Haematol., 112, 594-606,1985. kindreds. lO.GIROLAMI, A., SIMIONI, P., LAZZARO, A.R., CORDIANO, I. Severe arterial cerebral thrombosis in a patient with protein S deficiency (moderately reduced total and markedly reduced free protein S):a family study. Thromb. Haemost., 61, 144-147,1989. ll.LAURELL, phoresis 1968.
C.B. Quantitative estimation of proteins of human serum proteins. Anal. Biochem.,
by electro35, 403-413,
30
CO-FACTOR II DEFICIENCY
Vol. 67, No. 1
BOYER-NEUMANN, C., MARTINOLI, J.L., LEROY-MATHERON, 12.WOLF, M., J., MEYER, D., LARRIEU, M.J. A new functional C ., AMIRAL, assay for human protein S activity using activated factor V as substrate. Thromb. Haemost., 62, 1144-1145, 1989. 13.EGEBERG, 0. Inherited antithrombin III deficiency causing Thromb. Diath. Hemorrh., 13, 516-530, 1965. thrombophilia. 14.GRIFFIN, WIEDEMAN, disease.
J.H., EVATT, B., ZIMMERMANN, T.S., KLEISS, A.J., C. Deficiency of protein C in congenital thrombotic J. Clin. Invest., 68, 1370-1373, 1981.
L., MULLER, 15.BERTINA, R.M., VAN DER LINDEN, I.K., ENGESSER, E.J.P. Hereditary heparin cofactor II H.P., BROMMER, deficiency and the risk of development of thrombosis. Thromb.Haemost., 57, 196-200, 1987. 16.GIROLAM1, A., CAPPELLATO, M.G., VICARIOTO, M.A., CASONATO, S., F. Associated von Willebrand disease as a possible MARAFIOTI, cause of lack of thrombosis in an AT III abnormality (AT III Trento). Blut, 52, 29-33, 1986. 17.TAKEUCH1, Y., SAITO, Y., IKENOUCHI, H., SUGIMOTO, T., SUGOH, T ., SAKATA, Y., MATSUDA, M., TAKAMIYA, 0. Combined factor VII and protein C deficiency with peripheral pulmonary artery stenosis accompanied by progressive pulmonary hypertension and hemoptysis. Thromb. Res., 51, 117-126, 1988. 18.GRANDILLE, S., PRIOLLET, P., CAPRON, L., RONCATO, M., of inherited FIESSINGER, J.N., AIACH, M. Association dysfibrinogenaemia and protein C deficiency in two unrelated families. Br. J. Haemat., 68, 329-337, 1988. lg.MANABE, S., MATSUDA, M. Homozygous protein C deficiency combined with heterozygous dysplasminogenemia found in a 21 -year-old thrombophilic male. Thromb. Res., 39, 333-341, 1985. 2O.MAURIN, N., SCHUNKERT, H., combination of antithrombin Thromb. Haemost., 60, 127,
SIEBERTH, H.G. Thrombophilia with III and factor XII deficiency. 1988.
a
21.BRENNER, B., TAVORI, S., ZIVELIN, A., KELLER, C.B., SUTTIE, I., SELIGSOHN, J.W., TATARSKY, U. Hereditary deficiency of all vitamin K-dependent procoagulants and anticoagulants. Br. J. Haemat., 75, 537-542, 1990.
