Use of transplant status codes to monitor access to kidney transplantation.

The kidney transplantation process involves a series of steps related to interest in transplantation, medical suitability, pretransplant workup, and eventual transplantation. We developed transplant status codes to monitor the progress of dialysis patients in Ohio, Kentucky, Indiana, and Illinois as they move through these steps. Of the first 2,137 dialysis patients monitored with these new codes, 29% were interested in receiving a transplant, 31% were medically suitable to receive a kidney transplant, 15% had completed a pretransplant workup, 13% were on a cadaveric transplant waiting list, and 2% were awaiting a living-related transplant. Examination of these steps by gender showed differences in interest (men 32% v women 26%, P =.01) and medical suitability (men 33% v women 28%, P =.01). The proportion of patients successfully completing each step differed across facilities. In conclusion, transplant status codes are a useful method to monitor access to transplantation. As part of a facility-specific quality improvement program, such codes may be used to develop targeted interventions to help patients move through the transplant process as expeditiously as possible.

Managing the lifeline: preemptive access management for better outcomes for hemodialysis patients and programs. Medical Review Board of The Renal Network, Inc.

Problems with vascular access for hemodialysis patients contribute substantially to the morbidity, mortality, and high costs associated with renal replacement therapy. Data from the Network 9/10 Regional Hemodialysis Vascular Access Quality Improvement Project show evidence of incremental improvements, with more native vein fistulae and fewer grafts. However, increased numbers of catheters were observed, and still fully 32% of the catheters in place were associated with no internal access created after more than 90 days on dialysis. This article reviews recent contributions to understanding patterns of access-related care, pathophysiology of vascular access complications, and new approaches to achieving the preferred native vein arteriovenous fistulae. A financial analysis shows that dialysis units that employ dedicated access management personnel can expect bottom-line benefits that will easily cover the added expense. These benefits will be in addition to improvements in morbidity and mortality for patients.

Interleukin-12 neutralization alters lung inflammation and leukocyte expression of CD80, CD86, and major histocompatibility complex class II in mice infected with Histoplasma capsulatum.

Histoplasma capsulatum induces a cell-mediated immune response in lungs and lymphoid organs of mammals. Resolution of primary infection in mice depends on interleukin-12 (IL-12), since neutralization of this monokine increases susceptibility to infection. The present study was designed to determine if blockade of IL-12 disrupts the protective immune response by altering the influx of lineage-specific cells into infected lungs and the numbers of cells expressing CD80, CD86, CD119, and major histocompatibility complex class II (MHC II) molecules. In mice given anti-IL-12, there was a 2.5-fold decrease in total numbers of T cells on days 3 to 10 of infection and a 4-fold increase in Mac-1/Gr-1(+) cells on days 7 and 10 compared to infected controls. CD80(+) lung cells from anti-IL-12-treated mice were 2- to 3-fold greater than those from controls on days 7 and 10, whereas the total numbers of CD86(+) cells were 2- to 3-fold less and MHC II(+) cells were 1.5- to 2-fold less on days 3 and 5. Cells expressing CD119 were reduced 1.5-fold on day 5. Treatment with monoclonal antibodies (MAb) to CD80, CD86, or both reduced the fungal burden slightly compared to that in rat immunoglobulin G-treated controls, whereas after IL-12 neutralization, blocking of CD80 reduced the tissue burden by 2. 5-fold and this correlated with a decrease in IL-4. Regardless, mortality was not altered by treatment with MAb to CD80 or CD86. We conclude that (i) IL-12 neutralization alters the nature of the inflammatory response in lungs and the expression of CD80 and CD86 on lineage-specific cells, (ii) the immune response during infection with H. capsulatum is controlled via mechanisms independent of the CD80 and CD86 costimulatory pathways, and (iii) decreased expression of CD86 and MHC II may modulate generation of optimal protective immunity.

Use of standardized ratios to examine variability in hemodialysis vascular access across facilities. Medical Review Board of The Renal Network, Inc.

The type of hemodialysis vascular access (catheter, fistula, graft) is an important determinant of patient morbidity and dialysis efficiency. The relative importance of patient versus provider factors in determining type of vascular access is unclear. We sought to develop a quality improvement tool that adjusts for differences in patient characteristics, thereby allowing examination of provider-related variability in types of vascular access used across facilities. We examined 15,339 patients from 216 chronic hemodialysis units in Indiana, Kentucky, Ohio, and Illinois and found that 20% of patients had catheters, 24% had fistulas, and 56% had grafts. Young, male, and white patients were more likely to have fistulas, whereas old, female, and black patients were more likely to have grafts. Diabetics were more likely to have catheters and less likely to have fistulas. New patients were more likely to have catheters and less likely to have grafts. A facility specific standardized catheter ratio (SCR), standardized fistula ratio (SFR), and standardized graft ratio (SGR) were calculated based on the actual number of patients with each type of vascular access divided by the expected number adjusted for patient characteristics. Facility SCRs ranged from 0.00 to 2.87. Of the 216 facilities, 38 (18%) had an SCR significantly less than 1.00, and 32 (15%) had an SCR significantly greater than 1.00. Similar variability was observed in SFRs and SGRs. In conclusion, the type of vascular access varies greatly across facilities. Use of standardized access ratios adjusted for patient characteristics may help providers examine processes of care that contribute to variability in access use. Analogous to the standardized mortality ratio, the SCR, SFR, and SGR should be effective quality improvement tools.

Vbeta4(+) T cells promote clearance of infection in murine pulmonary histoplasmosis.

T cells are essential for controlling infection with Histoplasma capsulatum. Because the T cell receptor is vital for transducing the biological activities of these cells, we sought to determine if exposure to this fungus induced an alteration in the Vbeta repertoire in lungs of C57BL/6 mice infected intranasally. Vbeta2(+) cells were elevated on day 3 after infection; Vbeta4(+) cells were higher than controls on days 7, 10, and 14 after infection. Vbeta10(+) cells were increased on days 14 and 21, and Vbeta11(+) exceeded controls only on day 14. We investigated the clonality and function of Vbeta4(+) cells because their expansion transpired during the critical time of infection, that is, when cellular immunity is activated. Sequence analysis demonstrated preferential use of a restricted set of sequences in the complementarity-determining region 3. Elimination of Vbeta4(+) cells from mice impaired their ability to resolve infection. In contrast, depletion of Vbeta7(+) cells, the abundance of which was similar to that of Vbeta4(+), did not alter elimination of the fungus. The identification of clonotypes of Vbeta4(+) cells suggests that a few antigenic determinants may drive proliferation of this subset, which is necessary for optimal clearance.

Evolution of the primary immune response to Histoplasma capsulatum in murine lung.

Histoplasma capsulatum induces a cell-mediated immune response in the lungs and lymphoid organs of mammals. In this study, we analyzed the progression of the cytokine and inflammatory reactions in the lungs of mice infected intranasally with H. capsulatum. We measured cytokine mRNA levels and determined the inflammatory cell populations during the active phase of infection (<3 weeks). Transcription of genes encoding interleukin-2 (IL-2), IL-4, and IL-12 and gamma interferon (IFN-gamma) was detectable as early as day 3 of infection, whereas a signal for IL-10 was never observed. Competitive PCR analysis demonstrated that enhanced expression of IL-12 mRNA was observed by day 3 and that expression of mRNA for IL-2 and IFN-gamma progressively increased from day 5 to day 10. All levels declined by day 14. Analysis of the inflammatory response revealed an initial elevation in myeloid cells (Mac-1+) and natural killer (NK) cells followed by a rise in T cells, predominantly CD4+ cells. Since IFN-gamma is a key factor in host defense, we performed cytoplasmic staining to determine the cell populations that produced this cytokine. The hierarchy of synthesis was CD4+ > CD8+ > NK cells. Thus, H. capsulatum provokes an orderly modulation of the inflammatory and cytokine responses in murine lungs.

A delta T-cell receptor deleting element transgenic reporter construct is rearranged in alpha beta but not gamma delta T-cell lineages.

T cells can be divided into two groups on the basis of the expression of either alpha beta or gamma delta T-cell receptors (TCRs). Because the TCR delta chain locus lies within the larger TCR alpha chain locus, control of the utilization of these two receptors is important in T-cell development, specifically for determination of T-cell type: rearrangement of the alpha locus results in deletion of the delta coding segments and commitment to the alpha beta lineage. In the developing thymus, a relative site-specific recombination occurs by which the TCR delta chain gene segments are deleted. This deletion removes all D delta, J delta, and C delta genes and occurs on both alleles. This delta deletional mechanism is evolutionarily conserved between mice and humans. Transgenic mice which contain the human delta deleting elements and as much internal TCR delta chain coding sequence as possible without allowing the formation of a complete delta chain gene were developed. Several transgenic lines showing recombinations between deleting elements within the transgene were developed. These lines demonstrate that utilization of the delta deleting elements occurs in alpha beta T cells of the spleen and thymus. These recombinations are rare in the gamma delta population, indicating that the machinery for utilization of delta deleting elements is functional in alpha beta T cells but absent in gamma delta T cells. Furthermore, a discrete population of early thymocytes containing delta deleting element recombinations but not V alpha-to-J alpha rearrangements has been identified. These data are consistent with a model in which delta deletion contributes to the implementation of a signal by which the TCR alpha chain locus is rearranged and expressed and thus becomes an alpha beta T cell.

Role of complement receptor type three and serum opsonins in the neutrophil response to yeast.

Previous studies have suggested that neutrophil complement receptor type three (CR3) has two binding sites: (1) a site for fixed iC3b that does not trigger ingestion or a superoxide (O2-) burst, and (2) a function-triggering site for the beta-glucan component of yeast (Saccharomyces cerevisiae) cell walls. In the present study it was found that yeast (Y) coated with C3b (YC3b) or iC3b (YC3bi), prepared with purified complement in an IgG-free system, were avidly ingested ans stimulated a vigorous O2- burst, whereas sheep erythrocytes (E) bearing C3b or iC3b, were not ingested and did not give an O2- burst. YC3b and YC3bi contained an amount of fixed C3 that was approximately equal to serum-opsonized Y (OY), and produced O2- bursts comparable to OY. Experiments utilizing rabbit F(ab')2 anticomplement receptor type one (anti-CR1) to block fixed C3b binding to CR1, and monoclonal anti-CR3 (MN-41 or OKM1) to block fixed iC3b and Y cell wall binding to CR3, indicated that the O2- burst response to OY was primarily due to fixed iC3b and Y cell wall binding to CR3. Fixed C3b (that represented 33% of the fixed C3 on OY) and IgG anti-Y antibodies that bound to CR1 and Fc receptors, respectively, were found to contribute little to the response. Although YC3b did bind avidly to neutrophil CR1, the results suggested that the O2- burst response to YC3b was triggered after the initial YC3b binding by the secondary attachment of Y cell wall components to CR3. When neutrophils were treated with anti-CR3, 90% of neutrophils bound YC3b (via CR1), but phagocytosis and an O2- burst were completely absent. Similar findings were made with OKM1-treated neutrophils and YC3bi. Responses of OKM1-treated neutrophils were inhibited because only the iC3b-binding site of CR3 was ligated by the YC3bi. Thus, fixed C3b or iC3b on Y mediate avid binding of Y to neutrophils via CR1 or the iC3b-binding site of CR3, respectively, but ingestion and an O2- burst response are only triggered when glucans in the Y cell wall secondarily bind to neutrophils via the beta-glucan binding site of CR3.

Specificity of membrane complement receptor type three (CR3) for beta-glucans.

The binding of the iC3b receptor (CR3) to unopsonized zymosan was shown to result from CR3 attachment to cell wall beta-glucans. A specificity of neutrophil responses for beta-glucan was first suggested by a comparison of yeast (Saccharomyces cerevisiae) cell wall components for stimulation of a neutrophil superoxide burst. Neutrophils responded poorly to heat-killed yeast, but gave increasingly better responses to cell wall polysaccharides devoid of proteins (zymosan) and nearly pure beta-glucan particles derived from zymosan. Zymosan triggered a burst that was 29% as great as that stimulated by phorbol myristate acetate (PMA), and beta-glucan particles stimulated a burst that was 72% as great as that produced by PMA. Phagocytic responses to yeast were also inhibited by soluble glucans but not by soluble mannans. Three types of experiments demonstrated a role for CR3 in these responses. First, neutrophil ingestion of either yeast or yeast-derived beta-glucan particles was blocked by monoclonal anti-CR3, fluid-phase iC3b, or soluble beta-glucan from barley. Monocyte ingestion of beta-glucan particles was also blocked by anti-CR3, but not by anti-CR1 or anti-C3. Second, the neutrophil superoxide burst response to either zymosan or beta-glucan particles was blocked by anti-CR3 or fluid-phase iC3b, and was completely absent with neutrophils from 3 patients with an inherited deficiency of CR3. Third, CR3 was isolated from solubilized neutrophils by affinity chromatography on beta-glucan-Sepharose.

Membrane complement receptor type three (CR3) has lectin-like properties analogous to bovine conglutinin as functions as a receptor for zymosan and rabbit erythrocytes as well as a receptor for iC3b.

Human leukocyte complement receptor type three (CR3) was shown to be lectin-like and to resemble bovine serum conglutinin (K) in that it bound to both iC3b and unopsonized yeast (Saccharomyces cerevisiae), and was inhibited by EDTA or N-acetyl-D-glucosamine (NADG). CR3 and K also bound to zymosan (Z), a yeast cell wall extract that contains primarily polysaccharide and no detectable protein. However, structural differences and the absence of K on bovine phagocytes indicated that CR3 was not the human homologue of bovine K. Phagocytic and respiratory responses to unopsonized Z were CR3 dependent because they were inhibited by monoclonal antibodies specific for the alpha-chain of CR3 and did not occur with phagocytes from patients with a genetic deficiency of CR3. The binding of CR3 to Z did not require opsonization of the Z with neutrophil-secreted C3, as Z binding and responses were not inhibited by Fab anti-C3. In addition, CR3-dependent binding of yeast occurred with neutrophils from which protein secretion was blocked by fixation with paraformaldehyde. Rabbit erythrocytes (RaE) also bound weakly to neutrophil CR3 and triggered ingestion. Anti-CR3 not only blocked the binding and ingestion of RaE but also blocked selectively the ingestion of RaEC3b without affecting the strong binding mediated by CR1. Even though sheep E and sheep EC3b were not ingested by neutrophils, a weak binding of CR3 to sheep E was suggested by the finding of 20 to 40% inhibition of sheep EAIgG ingestion by anti-CR3. Such inhibition was only observed in buffers that allowed activity of the CR3 binding site and not in buffers containing either EDTA or NADG. An apparently contradictory finding was that the weak CR3-dependent binding of Z triggered neutrophil ingestion and a superoxide burst, whereas the avid CR3-dependent binding of sheep EC3bi did not induce significant ingestion or a respiratory burst. Blocking studies with monoclonal antibodies specific for different epitopes of the alpha-chain of CR3 suggested that this might result from the presence of two distinct binding sites in CR3: one site for fixed iC3b that did not trigger functions, and a second function-triggering site for Z that did not bind to fixed iC3b.

Generation of three different fragments of bound C3 with purified factor I or serum. II. Location of binding sites in the C3 fragments for factors B and H, complement receptors, and bovine conglutinin.

The many different recognized functions of C3 are dependent upon the ability of the activated C3 molecule both to bind covalently to protein and carbohydrate surfaces and to provide binding sites for as many as eleven different proteins. The location of the binding sites for six of these different proteins (factors B and H, complement receptors CR(1), CR(2) and CR(3) and conglutinin) was examined in the naturally occurring C3-fragments generated by C3 activation (C3b) and degradation by Factor I (iC3b, C3c, C3d,g) and trypsin (C3d). Evidence was obtained for at least four distinct binding sites in C3 for these six different C3 ligands. One binding site for B was detectable only in C3b, whereas a second binding site for H and CR(1) was detectable in both C3b and iC3b. The affinity of the binding site for H and CR(1) was charge dependent and considerably reduced in iC3b as compared to C3b. H binding to iC3b-coated sheep erythrocytes (EC3bi) was measurable only in low ionic strength buffer (4 mS). The finding that C3c-coated microspheres bound to CR(1), indicated that this second binding site was still intact in the C3c fragment. However, H binding to C3c was not examined. A third binding site in C3 for CR(2) was exposed in the d region by factor I cleavage of C3b into iC3b, and the activity of this site was unaffected by the further I cleavage of iC3b into C3d,g. Removal of the 8,000-dalton C3g fragment from C3d,g with trypsin forming C3d, resulted in reduced CR2 activity. However, because saturating amounts of monoclonal anti-C3g did not block the CR(2)-binding activity of EC3d,g, it appears unlikely that the g region of C3d,g or iC3b forms a part of the CR(2)-binding site. In addition, detergent-solubilized EC3d (C3d-OR) inhibited the CR(2)-binding activity of EC3d,g. Monocytes and neutrophils, that had been previously thought to lack CR(2) because of their inability to form EC3d rosettes, did bind EC3d,g containing greater than 5 x 10(4) C3d,g molecules per E. The finding that monocyte and neutrophil rosettes with EC3d,g were inhibited by C3d-OR, suggested that these phagocytic cells might indeed express very low numbers of CR(2), and that these CR(2) were detectable with EC3d,g and not with EC3d because C3d,g had a higher affinity for CR2 than did C3d. A fourth C3 binding site for CR(3) and conglutinin (K) was restricted to the iC3b fragment. Because of simultaneous attachment of iC3b to phagocyte CR3 and CR(3), the characteristics of iC3b binding to CR3 could only be examined with phagocytes on which the CR(1) had been blocked with anti-CR(1). Inhibition studies with EDTA and N-acetyl-D-glucosamine demonstrated a requirement for both calcium cations and carbohydrate in the binding of EC3bi to CR3 and to K. However, CR(3) differed from K in that magnesium cations were required in addition to calcium for maximum CR(3) binding activity, and NADG produced less inhibition of CR(3) activity than of K activity.