Neutrophil function and infections in uremia

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Manuel Martı´nez-Maldonado, MD Editor-at-Large

Neutrophil Function and Infections in Uremia Walter H. Ho¨rl, MD, PhD, FRCP

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NFECTIONS are frequent in uremic patients. Himmelfarb and Hakim1 summarized series, published between 1977 and 1989, reported a mortality rate from infections of 13.1% to 35.7% in these patients. In the 1997 US Renal Data report, septicemia caused death in 75% of the 15.5% of end-stage renal disease patients receiving renal replacement therapy who died of infection. Impairment of the host defense is primarily responsible for the susceptibility to infections. MALNUTRITION IMPAIRS CELL-MEDIATED IMMUNITY

A substantial number of end-stage renal disease patients develop malnutrition as a result of reduced protein and caloric intake. Loss of vitamins, trace elements, and micronutrients during dialysis therapy contribute not only to malnutrition, but also to impaired cell-mediated immunity and to the enhanced risk for infection in these patients. IRON OVERLOAD AND THE RISK FOR INFECTION

Iron overload is associated with leukocyte abnormalities and a greater frequency of infectious complications. The dysfunction of polymorphonuclear leukocytes (PMNLs) in iron-overloaded hemodialysis patients is partly related to the accumulation of iron in neutrophils. Therapy with recombinant human erythropoietin (rHuEPO) decreased serum ferritin levels and improved impaired phagocytosis. Iron deficiency leads to hyporesponsiveness to rHuEPO in patients with end-stage renal disease. High-dose iron therapy to avoid this may enhance the risk for iron overload and a greater frequency of infectious complications. In a recent study,2 healthy subjects were compared with

hemodialysis patients treated with rHuEPO and iron saccharate, iron-overloaded primary hematologic patients, and patients with hereditary hemochromatosis. Compared with PMNLs obtained from healthy subjects, those from hemodialysis patients showed mild inhibition of phagocytosis but significant inhibition of intracellular killing of bacteria. The in vitro oxidative burst of PMNLs was also significantly reduced in some dialysis patients. The PMNL defect of dialysis patients with high ferritin was similar to that of patients with exogenous or endogenous (hemochromatosis) iron overload, suggesting a direct inhibitory effect of iron on PMNLs.2 IMPAIRMENT OF PMNL FUNCTION BY INCREASED CYTOSOLIC CALCIUM

Secondary hyperparathyroidism is responsible for the elevation of cytosolic calcium in uremia. Increased cellular calcium [Ca2⫹ ] i inhibits mitochondrial oxygen consumption and oxidative phosphorylation with a consequent decrease in adenosine triphosphate (ATP) production. Reduction in ATP impairs the function of calcium pumps and reduces the extrusion of calcium out of the PMNLs, resulting in a sustained increase in [Ca2⫹] i. This process continues until a steady state with higher resting levels of [Ca2⫹] i and lower ATP content is achieved. As a consequence, PMNLs show impaired phagocytosis, decreased glucose uptake, inhibition of glycogen metabolism, and decreased oxygen consumption. In one study,3 normalization of cytosolic calcium by treatment with calcium channel blockers resulted in improved PMNL function independent of the plasma parathyroid hormone level. Decreasing parathyroid hormone level by 1,25

American Journal of Kidney Diseases, Vol 33, No 2 (February), 1999: pp xlv-xlviii

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dihydroxyvitamin D3 therapy also resulted in normalization of cytosolic calcium and improvement of the glucose uptake of PMNLs isolated from hemodialysis patients. IMPAIRED CELLULAR HOST DEFENSE BY LOW- AND HIGH-MOLECULAR-WEIGHT UREMIC TOXINS

A granulocyte inhibitory protein (GIP I) that is responsible for PMNL dysfunction in uremia has been isolated and characterized. This polypeptide inhibits the uptake of deoxyglucose, oxidative metabolism, chemotaxis, and intracellular bacterial killing by PMNLs.4 GIP I shares homol-

Walter H. Ho¨rl, MD, PhD, FRCP Head, Department of Nephrology University of Vienna, Austria

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HE ALLGEMEINES Krankenhaus, Vienna’s university hospital, is an enormous health complex that houses 2,200 hospital beds. If it is true that its lobby feels more like a cavernous railway station, it is equally true that some of the best nephrology in Europe is practiced there. In 1992, Walter H. Ho¨rl, already a distinguished nephrologist in his native Germany, became the head of the nephrology department at the University of Vienna. Ho¨rl completed his medical career in Wu¨rzburg, where he also trained in biology and biochemistry. Exposure to these disciplines ignited the imagination of the young physician and, immediately on graduation, sent him to the department of physiological chemistry at the University of Bochum, a city in the Ruhr basin not far from where Germany borders with Belgium and Holland. On receiving his PhD, he returned to Wu¨rzburg to complete his clinical fellowship before moving to the University of Freiburg as an Associate Professor of Medicine in 1983. Ho¨rl had already developed a keen interest in the mechanisms and pathophysiology of the uremic process, and his publications on this and other subjects made their mark. In 1990, Ho¨rl became the head of the department of nephrology at the University of Homburg in the Saar. The investigations he had begun in Wu¨rzburg on neutrophil metabolism in the uremic environment continued to advance toward the isolation of specific

ogy with light chain proteins. A second granulocyte inhibitory protein (GIP II), isolated from plasma ultrafiltrates from hemodialysis patients, also interferes with PMNL functions. GIP II shows homology with ␤2-microglobulin and inhibits in vitro oxygen production by PMNLs, as well as glucose uptake stimulated by phorbol ester. Intact ␤2-microglobulin does not influence these granulocyte functions. In contrast, GIP I inhibits formyl methionyl leucyl phenyalanine (FMLP)-mediated PMNL functions. GIP II stimulates interleukin-1␤ and interleukin-6 production by cultured human mononuclear cells. This effect is similar to that of ␤2-microglobulin modisubstances that impede the function of those cells. He discusses this topic in the accompanying article.In addition, he and his collaborators produced an impressive body of work on the effects of erythropoietin therapy on cardiovascular function and metabolism, the consequences of membrane biocompatibility in hemodialysis, the outcomes of continuous ambulatory peritoneal dialysis, experimental and clinical aspects of acute renal failure, and the pathophysiology of hypertension in uremia. Since arriving in Vienna in 1992, Ho¨rl has continued his astounding productivity; to date, his publications exceed 300 articles in international journals and more than 30 chapters in textbooks! For these contributions, as well as his clinical acumen and dedication to patients, he has been elected to the Royal College of Physicians (London) and was recently chosen by his peers as one of the top nephrologists in Europe. A member of numerous editorial boards, including the American Journal of Kidney Diseases, he is also one of the editors of new German textbooks of internal medicine and blood purification. Walter Ho¨rl is a world-known nephrologist and clinical scientist, but he is even better known for his generosity toward his colleagues, particularly those who are young and in their developmental phase. A man of many interests and talents, modesty, and easy charm, he admits—from behind his characteristic grin—that the best decision he has made in his awesome career was to marry his wife Ursula. Ursula, a watercolorist and a veritable walking encyclopedia of the art, architecture, and history of the Austro-Hungarian Empire (among many, many other things), with a twinkle in her eye bemoans her slight knowledge of nephrology. Professor and Mrs Ho¨rl have three very lucky sons. —Manuel Martı´nez-Maldonado, MD

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fied by advanced glycation end products. Intact ␤2-microglobulin was without effect on cytokine production. Both GIP I and II were also isolated and characterized from peritoneal effluents (of continuous ambulatory peritoneal dialysis [CAPD] patients) that inhibited the PMNL properties. A PMNL degranulation inhibiting protein (DIP I) was purified from plasma ultrafiltrates obtained from patients undergoing regular hemodialysis therapy. DIP I is identical to angiogenin and inhibits spontaneous, as well as stimulated, PMNL degranulation.5 A polyclonal antibody to human recombinant angiogenin abolishes the inhibitory effect of the isolated protein on PMNL. A similar effect is induced by the disulfide C39 to C92–containing tryptic angiogenin fragment, indicating a new, biologically active site of angiogenin that is different from the sites responsible for the angiogenetic activity of the protein. Plasma angiogenin levels are significantly elevated in hemodialysis and CAPD patients; thus, it may have a role in PMNL degranulation. A second degranulation inhibiting protein (DIP II) was isolated from human plasma ultrafiltrate and identified as complement factor D by means of sequence analysis. Complement factor D caused a dose-dependent decrease of stimulated lactoferrin degranulation to 34% of stimulated controls. Serum of healthy donors contains low concentrations of factor D (1 to 2 µg/mL), whereas this level can be increased tenfold in the serum of dialysis patients. Blood ultrafiltrates and peritoneal effluents of patients undergoing hemodialysis or peritoneal dialysis contain kappa and lambda light chains. Nanomolar concentrations of these proteins inhibit in vitro glucose uptake and chemotaxis of PMNLs.6 Usually, a small amount of light chains exist in free form in the serum, but up to a fivefold increase in the level of free immunoglobulin light chains has been shown in sera from patients with severely reduced kidney function. A peptide that inhibits the chemotactic movement of polymorphonuclear leukocytes in a concentration-dependent, nonreversible manner has been isolated from the peritoneal effluent of peritoneal dialysis patients.7 The isolated peptide has a more acidic isoelectric point but the same amino terminal sequence as ubiquitin and is

xlvii Table 1. Granulocyte Inhibitory Proteins Isolated From Patients on Hemodialysis and CAPD 1. Granulocyte inhibitory protein I (80% homology to the kappa and 40% homology to the lambda light chain sequence; MW, 28,000 d) 2. Granulocyte inhibitory protein II (homology to ␤2microglobulin; MW, 9,500 d) 3. Degranulation inhibiting protein I (identical to angiogenin; MW, 14,400 d) 4. Degranulation inhibiting protein II (identical to complement factor D; MW, 24,000 d) 5. Immunoglobulin light chains (kappa mono- and dimers, lambda mono- and dimers) 6. Chemotaxis inhibiting protein (modified ubiquitin; MW, 8,500 d) Abbreviation: MW, molecular weight.

identified by antiubiquitin antibodies in Western blot experiments. Antiubiquitin antibody binding fractions were isolated from all peritoneal dialysis and hemodialysis patients investigated. These fractions, containing the same acidic band, also significantly inhibited PMNL chemotaxis. Ubiquitin per se had no effect on PMNL chemotaxis, leading to the suggestion that a modified form of ubiquitin was isolated and that this modification was responsible for its inhibitory effect. Table 1 lists the GIPs isolated and characterized so far from high-flux dialyzer ultrafiltrates and peritoneal effluents of uremic patients. CONCLUSION

Chronic uremia, particularly if associated with malnutrition, results in profound alterations of the immune system. PMNL functions, such as chemotaxis, oxidative metabolism, phagocytic activity, degranulation, intracellular killing, or carbohydrate metabolism, are inhibited by lowand high-molecular-weight inhibitors. Iron overload secondary to intravenous iron treatment of the anemia of end-stage renal disease and the accumulation of intracellular calcium in patients with secondary hyperparathyroidism results in deactivation of neutrophils. REFERENCES 1. Himmelfarb J, Hakim RM: Biocompatibility and risk of infection in haemodialysis patients. Nephrol Dial Transplant 9:138-144, 1994 (suppl 2) 2. Patruta SI, Edlinger R, Sunder-Plassmann G, Ho¨rl WH: Neutrophil impairment associated with iron therapy in hemodialysis patients with functional iron deficiency. J Am Soc Nephrol 9:655-663, 1998

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3. Ho¨rl WH, Haag-Weber M, Mai B, Massry SG: Verapamil reverses abnormal [Ca2⫹ ] i and carbohydrate metabolism of PMNL of dialysis patients. Kidney Int 47:1741-1745, 1995 4. Ho¨rl WH, Haag-Weber M, Georgopoulos A, Block LH: Physicochemical characterization of a polypeptide present in uremic serum that inhibits the biological activity of polymorphonuclear cells. Proc Natl Acad Sci U S A 87:63536357, 1990 5. Tschesche H, Kopp C, Ho¨rl WH, Hempelmann U:

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Inhibition of degranulation of polymorphonuclear leukocytes by angiogenin and its tryptic fragment. J Biol Chem 269:30274-30280, 1994 6. Cohen G, Haag-Weber M, Mai B, Deicher R, Ho¨rl WH: Effect of immunoglobulin light chains from hemodialysis and CAPD patients on PMNL functions. J Am Soc Nephrol 6:1592-1599, 1995 7. Cohen G, Rudnicki M, Ho¨rl WH: Isolation of modified ubiquitin as a neutrophil chemotaxis inhibitor from uremic patients. J Am Soc Nephrol 9:451-456, 1998