International Urology and Nephrology 27 (5), pp. 643-647 (1995)

Haemostatic Parameters in Childhood Nephrotic Syndrome (Is there Any Difference in Protein C Levels between Steroid Sensitive and Resistant Groups?) F, YAL(~INKAYA,* N. TOMER,* A. N. GORGANI,** M. EKIM,* N. ~AKAR* Departments of *Paediatric Nephrology, Ankara University Faculty of Medicine; **Biochemistry, Hacettepe University Faculty of Medicine, Ankara, Turkey (Accepted January 23, 1995) Plasma protein C (PC) activity, prothrombin time (PT), partial thromboplastin time (PTT) and platelet count were studied in 23 children with nephrotic syndrome (NS) and AT III activity was determined in 15 children in the same group. All parameters were compared with those obtained in a group of 16 age matched healthy controls. Mean plasma AT III activity was significantly reduced in patients with NS (P <0.001 ) correspondingly, plasma AT III levels were found to be directly correlated with serum albumin and inversely correlated with proteinuria. In contrast, mean plasma PC activity, as well as PT, PTF and platelet count were similar in the NS group when compared with the control group and in addition no remarkable difference was found in the mean plasma PC activity between the steroid sensitive and resistant NS groups. In conclusion, this study demonstrated acquired AT III deficiency and normal PC levels in childhood NS. These data suggest that although plasma AT III activity depends on the severity of NS, neither the severity of NS nor the underlying renal disease is an important factor determining the changes of PC activity in childhood NS.

Patients with nephrotic syndrome (NS) have an increased tendency to thrombotic complications. A number of abnormalities of the haemostatic mechanism have been described in the pathogenesis of clotting diathesis, including elevated fibrinogen, cofactor V and VIII levels, reduced fibrinolysis, thrombocytosis and increased platelet aggregation [ 1-5]. None of these abnormalities, however, has been demonstrated consistently in nephrotic patients. Many studies have documented antithrombin III (AT III) deficiency [6-8], but data on changes of plasma protein C (PC) levels are limited, especially in childhood NS [7, 9, 10]. PC is a vitamin K dependent natural anticoagulant and its activity has been shown to be normal [11-13] or increased [7, 9, 10, 14] in patients with NS. The present study was undertaken to examine some haemostatic parameters including prothrombin time (PT), partial thromboplastin time (PTT), platelet count, PC and AT III levels in childhood NS and to investigate whether there is any difference in PC levels between steroid sensitive and resistant groups.

VSP, Utrecht Akad~miaiKiad6, Budapest

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Yalfinkaya et al.: Nephrotic syndrome

Patients and methods

The study included 23 patients (16 boys, 7 girls) with NS. Their ages ranged from 2.5 to 13 years with a mean of 8.57+3.72 years. NS was defined as heavy proteinuria (>40 mg/h m 2) and hypoalbuminaemia (<3 g/dl) with or without oedema. All of the patients had normal renal and liver function and none of them were receiving drugs. Patients with primary eoagulopathies or acute thrombotic events were excluded from the study. Thirteen patients were steroid sensitive and responded to a daily course of steroid therapy -dithin four weeks. Ten patients who failed to respond had renal biopsies. The underlying renal disease of the steroid resistant patients included renal amyloidosis (4), focal glomerulosclerosis (2), IgM mesangial proliferation (1), lupus nephritis (1), membranous nephropathy (1), membranoproliferative glomerulonephritis (1). PT, PTT, platelet count and plasma PC activity were determined in all patients and AT III activity was measured in 15 patients. Sixteen age matched healthy children (3 boys, 13 girls) served as a control group. Blood samples for assays were anticoagulated with sodium citrate in a ratio of 9:1. Platelet poor plasma was obtained by centrifugation at 3000 g for 15 min, stored at -70 ~ and tested within four weeks. The plasma levels of PC and AT III were determined by a chromogenic technique using commercial kits (Stachrom PC and AT III from Diagnostica Stago, Paris). Serum creatinine, albumin, ALT, AST levels, urinary protein excretion, PT, PTT and platelet count were determined routinely. The results were expressed as mean+standard deviation. Student's t-test and the Mann-Whimey test were performed to assess the significance of the differences between the groups. Correlation analysis was done by linear regression. Results

The results are shown in Table 1. Plasma PC activity, PT, PTT and platelet counts were similar in both groups (P>0.05). Plasma AT III activity was significantly lower (P<0.001) in the NS group (62.33+19.74%) than that in the control group (98.5+18.04%). There was a significant positive correlation (r = +0.807, P<0.001) between the AT III activity and serum albumin and in addition a significant negative correlation (r = -0.77, P<0.001) was noted between this concentration and urinary protein excretion. PC activity did not correlate with either serum albumin levels (r = +0.09, P>0.05). Protein C and AT III activity also did not correlate with each other (r = -0.01, P>0.05). Figure 1 shows the PC levels in patients with steroid sensitive and resistant NS. There was no significant difference (P>0.05) in the mean level of PC activity between steroid sensitive (92.92+26.99%) and resistant (83.2411.86%) NS groups. AT III activity was determined in 15 patients (thirteen of them were steroid sensitive NS). As the patient population was small, statistical analysis was not available for AT III activity in steroid sensitive and resistant groups. International Urology and Nephrology27, 1995

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Table 1 Summary of the results of haemostatic parameters in patients with NS and the control group NS group Serum albumin (g/dl) PC** (%)

AT III* (%) PT** (s) PTT** (s) Platelet count/mm3

Controlgroup

1.83+0.51 (n=23) 88.35 +21,8 (n=23) 62,334-19.74 (n=15) 14,09+1.24 (n=23) 4139+7.04 (n=23) 282.6964-90.44 (n=23)

4.774-0.44

(n=16) 98.88+19.90 (n= 16) 98.5+18.04 (n=16) 14.9+1,31 (n= 16) 46.624-9.06 (n= 16) 236.6764-77.41 (n= 16)

* PO,05 Steroid sensitive NS

Steroid resistant NS

140 120OQ

o~ 100-

|

!

80-

.% 6040-

Fig. 1. Plasma PC levels m patients with steroid sensitive and resistant NS

Discussion

The acquired AT III deficiency observed in our patients confhans the previous reports [6-8]. Correspondingly, plasma AT III levels were found to be positively correlated with serum albumin and inversely correlated with proteinuria. These results suggest that plasma AT III activity depends on the severity o f NS in childhood. International Urologyand Nephrology 27, 1995

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Our study revealed that mean plasma PC activity in children with NS was not statistically different from that of the control group. The conflicting resuits reported in previous studies may be due to differences in clinical material [7, 9, 10-14]. Our group consisted of children only and none of them had renal failure or were receiving drugs that may alter haemostasis. Moreover, most studies on haemostasis in nephrosis have examined heterogeneous groups of patients with NS without considering the underlying renal disease [8, 9, 14]. In the present study no remarkable difference was found in mean plasma PC activity between the steroid sensitive and resistant NS groups. This supports the observations reported by Vaziri et al. [14] and Mannucci et al. [10] who found no discernible relation between the underlying renal disease and PC levels. But, our results vary from those of Allan et al. [15] who observed a marked elevation of PC antigen in focal glomerulosclerosis and membranous glomerulonephritis. In conclusion, our study demonstrated that AT III activity is decreased and PC activity is normal in childhood NS. As the low level of AT III is correlated with the severity of NS, acquired AT III deficiency may favour a thrombotic tendency in NS. It was also shown that neither the severity of NS nor the underlying renal disease (steroid sensitive or resistant) is an important factor determining the changes of PC activity in children.

References 1. Bernard, D. B.: Extrarenal complications of the nephrotic syndrome. Kidney Int., 33, 1184 (1988). 2. Cameron, J. S.: The nephrotic syndrome and its complications. Am. J. Kidney Dis., 10, 157 (1987). 3. Kendall, A. G., Lehmann, R. C., Dossetor, J. B.: Nephrotic syndrome. A hypercoagulable state. Arch. lntern. Med., 127, 1021 (1971). 4. Llach, F.: Hypercoagulability, renal vein thrombosis and other thrombotic complications ofnephrotic syndrome. Kidney Int., 28, 429 (1985). 5. Massry, S. G., Vaziri, N. D.: Renal vein thrombosis and the nephrotic syndrome. In: Massry, S. G., Glassock, R. J. (eds): Textbook of Nephrology. Williams and Wilkins, Baltimore 1989. 6. Kauffmann, R. H., Veltkamp, J. J., Tilburg, N. H. V.: Acquired antithrombin III deficiency and thrombosis in the nephrotic syndrome. Am. J. Med., 65, 607 (1978). 7. Ueda, N,: Effect of corticosteroids on some haemostatic parameters in children with minimal change nephrotic syndrome. Nephron, 56, 374 (1980). 8. Vaziri, N. D., Panle, P., Toohey, J.: Acquired deficiency and urinary excretion of antithrombin III in nephrotic syndrome. Arch. lntern. Med., 144, 1802 (1984). 9. Pabinger-Fasching, I., Lechner, K., Niessner, H.: High levels of plasma protein C in nephrotic syndrome. Thromb. Haemostas., 53, 3 (1985). 10. Mannucci, P. M., Valsecchi, C., Bottasso, B.: High plasma levels of protein C activity and antigen in the nephrotic syndrome. Thromb. Haemostas., 55, 31 (1986). 11. Sala, N., Oliver, A., Estivill, X.: Plasma and urinary protein C levels in nephrotic syndrome. Thromb. Haemostas., 54, 900 (1985). 12. Soff, G. A., Sica, D. A., Marlar, R. A.: Protein C levels in nephrotic syndrome: Use of a new enzyme-linked immunoadsorbent assay for protein C antigen. Am. J. Hematol., 22, 43 (1986). International Urologyand Nephrology27, 1995

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13. Sorensen, P. J., Knudsen, F., Nielsen, A. H.: Protein C activity in renal disease. Thromb. Res., 55, 243 (1985). 14. Vaziri, N. D., Alikhani, S., Patel, B.: Increased levels of protein C activity, protein C concentration, total and free protein S in nephrotic syndrome. Nephron, 49, 20 (1988). 15. Allan, M., Softer, O., Evatt, B. L.: Protein S and C antigen levels in proteinuric patients: Dependence on type of glomerular pathology. Am. J. Hematol., 31, 96 (1989).

International Urologyand Nephrology27, 1995