Harald Lange

Noninvasive monitoring using serum amyloid A and serum neopterin in cardiac transplantation

The monitoring of allograft function for cardiac transplant patients still relies on endomyocardial routine biopsies. We investigated the diagnostic value of noninvasive monitoring using the parameters serum amyloid A protein and serum neopterin. The circulating levels of the acute phase reactant, amyloid A protein, and the macrophage product, neopterin, were measured serially in 13 patients after cardiac transplantation. The mean period of observation was 240 days. Nine acute cardiac allograft rejections, five cases of viral infection and eight cases of bacterial infection occurred. The levels of serum amyloid A protein and serum neopterin remained low (x = 6.0 mg/dL and 12.6 nmol/L, respectively) during the periods of stable graft function. In contrast, both parameters were significantly elevated (p < 0.01) during the rejection episodes (x = 12.7 mg/dL and 38.0 nmol/L for serum amyloid A protein and serum neopterin, respectively). For a reliable differentiation between rejection and stable graft function, serum amyloid A protein had a diagnostic accuracy of 84% (with a cut-off level of 10 mg/dL) and serum neopterin had one of 75% (with a cut-off level of 23 nmol/L). However, significant increases in the circulating levels of serum amyloid A protein and serum neopterin were also observed during bacterial (x = 14.9 and 88 nmol/L, respectively) and viral (x = 6.2 mg/dL and 44 nmol/L, respectively) infections. The detection of immunological complications after cardiac transplantation using serial measurements of serum amyloid A protein and serum neopterin is possible. These parameters can be used to help in judging both the need and the optimal timing for the otherwise frequent endomyocardial biopsies.

Detection of renal allograft rejection by complement components C5A and TCC in plasma and urine

Allograft rejection is associated with complement activation. Yet inconsistent results were obtained in evaluating plasma levels of complement factors or activation products as rejection markers. Therefore the human anaphylatoxin C5a and the soluble terminal complement complex (TCC) were measured by daily enzyme immunoassays on plasma (P) and urine (U) samples from 28 patients undergoing renal transplantation over a mean postoperative period of 25.8 days. The complement levels were evaluated longitudinally (cutoff of 100% increase on the previous days level) during periods of rejection, stable graft function, acute tubular necrosis, and cytomegalovirus disease. Regarding the detection of 13 acute rejection episodes, U-C5a showed a diagnostic accuracy of 81% (sensitivity of 85%, specificity of 77%), P-C5a one of 62%, and P-TCC one of only 30%. The U-C5a increment (mean rise of 379%) preceded the clinical diagnosis of rejection by an average of 1.6 days. Cytomegalovirus diseases (n = 4) were associated with high P-C5a levels (mean increase of 251% by the time of the first detection of viral DNA). In contrast, resumption of kidney function after acute tubular necrosis (n = 10 periods) was heralded by marked peaks of U-C5a (x = 43.7 microg/l). U-TCC was not detected in any clinical setting. In conclusion, as opposed to P-TCC, U-TCC, and P-C5a, the anaphylatoxin C5a, measured daily in urine, might have potential as an early and reliable marker for acute renal allograft rejection.

Biocompatibility Differences with Respect to the Dialyzer Sterilization Method

The impact of the method of sterilization (steam vs. ethylene oxide, ETO) on indices of biocompatibility is investigated using polysulfone membranes. Eight patients were treated with a random choice of the high-flux membranes F60S (steam) and F60 (ETO) and the low-flux membrane F6 (ETO). Blood samples were taken prior to and 5, 15, 30, 60, and 180 min after the start of hemodialysis. White blood cell count, platelet count, and plasma concentrations of polymorphonuclear neutrophil elastase, complements C3a and C5a, and β2-microglobulin were determined. The dialysis procedure was associated with a significant decrease in white blood cell count and β2-microglobulin level and a significant increase in polymorphonuclear neutrophil elastase and complement C3a and C5a levels. However, the steam-sterilized F60S membrane had a significantly lower impact on the biocompatibility indices than the ETO-sterilized F60 and F6 membranes (p < 0.05 or p < 0.001 for the individual markers). We conclude that using steam instead of ETO for sterilization may improve the biocompatibility of membranes.

Pattern and persistence of the epitope-specific IgM response against human cytomegalovirus in renal transplant patients

The humoral immune response against human cytomegalovirus (HCMV) was evaluated in immunocompromised patients by Western blotting (WB) based on recombinant viral envelope (gB and gH) and tegument (pp150 and pp65) proteins. Three groups of patients were investigated: (a) 74 renal transplant recipients; (b) 24 hemodialysis patients, both groups without clinical evidence of viral infections; and (c) 19 renal transplant patients with manifest HCMV infections. The results obtained suggest that (i) the WB is considerably more sensitive, recognizing the HCMV-specific IgM response rather than the enzyme-linked immunosorbent assays. An IgM response was detected in one-third of all clinically asymptomatic renal patients. (ii) The virus-specific IgM response is primarily directed against the pp150 epitope. (iii) In patients with clinically manifest HCMV disease, additional IgM reactivities are most frequently directed against the glycoprotein B epitope. (iv) The severity of HCMV infections correlates with the extent of the IgM antibody response, i.e. with the number of specific epitopes involved. (v) After transplantation, IgM reactivity and its epitope-specific pattern persist for years.

Impact of Profile Haemodialysis on Intra-/Extracellular Fluid Shifts and the Release of Vasoactive Hormones in Elderly Patients on Regular Dialysis Treatment

In 15 patients with end-stage renal failure and proven coronary heart disease, profile haemodialysis with decreasing ultrafiltration rate and hyperionic, decreasing dialysate solute concentration was compared with conventional, extracorporeal bicarbonate haemodialysis (Na+D = 138 mmol/l). Body fluid distribution and the release of vasoactive hormones (plasma renin activity, aldosterone, norepinephrine, epinephrine, and atrial natriuretic peptide) were investigated. Haemodialysis with constant ultrafiltration rate and constant dialysate composition (A) was followed by two dialysis profiles: decreasing ultrafiltration rate (B) and additional hyperionic, decreasing dialysate sodium concentration (C). In all 15 patients, the dialysis procedures (A) - (C) were used for 2 weeks each with six sessions, the last being taken for investigation. Body fluid distribution was calculated. In patients with serum sodium above 136 mmol/l, the conventional dialysis (A) as well as the Uf profile (B) showed a net fluid shift from extracellular volume (ECV) to intracellular volume (ICV). Using the profile with hyperionic, decreasing Na+D (C), the reverse fluid shift with decreasing ICV was achieved not only in those with serum Na+ <136 mmol/l, but also in those with serum Na+ > or = 136 mmol/l. The release of vasoactive hormones decreased already at profile haemodialysis (B) compared with (A) and was further reduced in (C). These results would suggest, profile dialyses B and C to have less impact on the cardiovascular system in elderly patients assuming higher patient comfort compared with the standard dialysis procedure. A higher benefit was obtained in C compared with B, presumably due to the additional prevention of the ICV shift and plasma volume depletion in patients with initial serum sodium > or = 136 mmol/l using transiently hyperionic Na+D. These results show that in elderly patients, hyperionic profile haemodialysis (Na+D > Na+S) had less impact on cardiovascular regulation than conventional bicarbonate dialysis.

Assessment of cytomegalovirus infections using neopterin and a new immunoblot

Human cytomegalovirus (HCMV) infections are a major cause of morbidity and mortality in immunocompromised patients despite advances in diagnostic tests and antiviral therapies. The underlying study investigates the diagnostic value of the immune marker neopterin and a recently developed HCMV-specific western blot to detect HCMV infections and to differentiate them into either syndromes or diseases. The mean period of observation was 1428 days. Thirteen HCMV diseases and nine syndromes were diagnosed retrospectively. The first appearance of clinical signs or symptoms was always associated with a marked increase of serum and urine neopterin. The HCMV-specific IgM response followed in the mean 9 days later. Median values and the course of the neopterin levels were significantly higher during the HCMV diseases. In addition, the strength of the humoral immune response was related to the severity of the HCMV infection. Patients with HCMV diseases developed antibodies against a higher number of epitopes. The anti-HCMV IgM response persisted in more than 80% of the patients for longer than 3 years. In conclusion, combining the HCMV-specific western blot and neopterin permit detection of the immune response against HCMV, reflect the severity of the infection and might guide the anti-viral therapy.

Hemodialysis and Blood Coagulation: The Effect of Hemodialysis on Coagulation Factor XIII and Thrombin-Antithrombin III Complex

Blood membrane interaction during hemodialysis (HD) regularly leads to stimulation of leukocyte function and related release of granular enzymes. The present study aimed to investigate the possible influence of an HD-induced release of granulocyte elastase on blood coagulation. Therefore a highly sensitive substrate of polymorphonuclear elastase, the plasma coagulation factor XIII and its subunits A and S were determined in the course of HD. Consumption of both subunit A and S have been previously shown to be due to proteolysis by elastase, whereas a decrease in subunit A will be typical for thrombin activation. Furthermore, the thrombin-antithrombin III complex (TAT) acting as a predisposition parameter for thrombotic events was measured during HD treatment. Apart from a virtual fall in factor XIII total activity simulated by heparin, no significant HD-induced consumption of factor XIII could be observed. There was also no indication of an elastase- or thrombin-related change in subunit concentrations. Predialysis values of the TAT complex were generally elevated in HD patients, but only patients with acute renal failure showed a constant increase of TAT during HD. These findings suggest that HD patients are exposed to a latent activation of coagulation resulting in an elevated thrombogenetic risk mainly due to the underlying disease. An additional coagulatory stimulation by the HD procedure seems to be restricted to cases of acute renal failure.

The anaphylatoxin C5a, a new parameter in the diagnosis of renal allograft rejection

Abstract  In the underlying study the diagnostic value of the anaphylatoxin CSa was evaluated in kidney transplantation. In 49 transplant patients the following parameters were measured daily for a mean period of 25.1 days: plasma a [P‐CSa], urine a [U‐CSa], serum amyloid A (SAA], serum neopterin [S‐NEOP] and urine neopterin [U‐NEOP]. Sensitivity, specificity and day of first significant parameter increase (exceeding a cut‐off level of > 50 YO) were evaluated retrospectively during 30 periods of rejection and 30 periods of stable graft function. U‐ a was the parameter with the highest sensitivity (84 %) and specificity (84 %), increasing in the mean 1.3 days before clinical diagnosis of rejection. Sensitivity and specificity of the other markers was lower: SAA 77 % and 77 %, U‐NEOP 68 % and 65 %, S‐NEOP 45 % and 77 %, and P‐ a 45 % and 48 %, respectively. During four instances of cy‐tomegalovirus disease extremely high U‐NEOP (≥= 1520 ± 518 μmol/mol creatinine) and slightly increased P‐ a levels (≥= 1.5 ± 1.4 ng/ml) occurred. Elevated urinary excretion of a seems to be a reliable and early marker of renal allograft rejection. In combination with SAA and U‐NEOP, the daily assessment of U‐ a differentiates between viral infection and allograft rejection.

Latent Inhibition of Granulocyte Function by Cyclosporine A

According to the literature, Cyclosporine A (CsA) is said to suppress specifically the activity of T and B cells. A significant influence on phagocyte function has been neglected. However, aggravated courses of bacterial and fungal infections have been frequently reported under the treatment with CsA, suggesting that a latent depression of phagocytic activity may possibly occur under clinical circumstances. Therefore, this study set out to assess whether CsA can also change granulocyte function under therapy conditions or not. Thirty-seven patients, 3 months-10 years after kidney transplantation being under immunosuppressive treatment with CsA + Prednisolone (n = 25), Azathioprine + Prednisolone (n = 6) and under Prednisolone alone (n = 6) underwent the study. 18 healthy persons served as a normal control group. Granulocyte function was tested ex vivo by chemiluminescence (CL) after stimulation with phorbolmyristate acetate (PMA) and with zymosan (zym) activated autologous or pool-serum. The obtained data were correlated to corresponding serum or plasma levels of CsA, human leukocyte elastase (HLE) and neopterin. Comparing the three therapy groups with the healthy control and with each other no differences could be seen in median CL values; but there was a significant (p = 0.05) negative correlation between CsA blood levels and maximum CL values of PMN. Such inhibition of CL could be calculated for zym but not for PMA stimulated PMN; suggesting that the CsA mediated inhibition of granulocyte function may be only partial and restricted to phagocytosis. In addition, a positive correlation between serum levels of human leukocyte elastase (HLE) and neopterin could be found. This indicates a simultaneous influence of CsA on both PMN and macrophages.

Kinetics of pulmonary leukocyte sequestration in man during hemodialysis with different membrane-types.

Although it has been suggested that pulmonary sequestration of leukocytes could account for membrane-dependent white blood cell depletion in HD, direct evidence in patients is still lacking. Therefore a study was initiated to test whether and how leukocytes distribute in the lung circulation during HD with different membranes. Thirteen patients suffering from chronic renal failure underwent lung scintigraphy during HD with cuprophane (n = 3), hemophane (n = 8) and polysulfone (n = 2) lowflux capillary dialyzers. Isolated autologous leukocytes were labelled with 99m-Technetium and reinfused before starting HD. Distribution of leukocyte related activity was registered by lung scintigraphy. In comparison to normal lung scintigraphy performed without HD, an impressive redistribution peak was demonstrated 10-20 min after the start of HD with cuprophane and also to a lesser extent with hemophane. When HD was performed with polysulfone the decrease in activity was delayed but no real redistribution was obtained. In accordance with other phenomena, such as peripheral leukopenia and changes in granulocyte oxidative metabolism, pulmonary sequestration of leukocytes takes place in man in the initial phase of HD and appears to be strongly dependent on the type of membrane. (Int J Artif Organs 1990; 13: 729-36)