Effectiveness of different immunoadsorption columns for anti-A/B antibody depletion.

OBJECTIVE: In recent studies, semi-selective compared to antigen-specific immunoadsorption (IA) columns showed comparable effectiveness in anti-A/B antibody removal before incompatible living donor kidney transplantation. Semi-selective columns allow a greater number of IA treatments at lower costs. They are also capable of removing potentially harmful human leukocyte antigen alloantibodies. Nevertheless, additional plasma exchange treatments are often necessary to reach the preoperative target titer, most likely due to an inadequate anti-A/B IgM antibody depletion. METHODS: We compared the effectiveness of immunoglobulin and anti-A/B antibody reduction by different semi-selective (Therasorb Ig-flex, Therasorb Ig-omni5, Immunosorba) and antigen-specific (Glycosorb) IA columns during the desensitization of 63 ABO-incompatible living donor kidney transplant candidates with a total of 375 IA treatments. Fifty-three patients were eventually transplanted. RESULTS: Total IgM reduction during the first IA treatment was significantly greater with the Therasorb Ig-omni5 compared to the Therasorb Ig-flex (mean: -71.3 vs -41.6; p = 0.001) or Immunosorba columns (mean: -71.3 vs -42.8; p = 0.03). During a median of 5.5-6 pre-transplant IA treatments, Therasorb Ig-flex and Therasorb Ig-omni5 columns were equally effective in the reduction of total IgM while both showed superior IgM reduction compared to the Immunosorba columns (Therasorb Ig-flex, mean: -81.2 vs -72.2; p = 0.01; Therasorb Ig-omni5, mean: -88.2 vs -72.2; p = 0.02). IgG reduction was not significantly different between groups. Likewise, anti-A/B IgM antibody reduction (titer Saline) during the first IA treatment was significantly greater with the Therasorb Ig-omni5 compared to the Therasorb Ig-flex (mean titer reduction: -1.9 vs -1.1; p = 0.02) and tended to be greater than with Immunosorba or Glycosorb columns. During a median of 5-6 pre-transplant IA treatments, overall anti-A/B IgM antibody reduction was significantly greater when IA was performed with the Therasorb Ig-flex (mean titer reduction: -3.8 vs -1.3; p < 0.001) or Therasorb Ig-omni5 (mean titer reduction: -4.3 vs -1.3; p = 0.01) compared to the Immunosorba columns with no differences compared to the Glycosorb columns. Again, anti-A/B IgG antibody reduction (titer Coombs) was not significantly different between groups. CONCLUSIONS: The semi-selective Therasorb Ig-omni5 device offers potential advantages in reducing total IgM as well as anti-A/B IgM antibodies.

Outcomes Following ABO-Incompatible Kidney Transplantation Performed After Desensitization by Nonantigen-Specific Immunoadsorption.

BACKGROUND: For desensitization of ABO-incompatible kidney transplant recipients we recently proposed nonantigen-specific immunoadsorption (IA) and rituximab. METHODS: We now compared clinical outcomes of 34 ABO-incompatible living-donor kidney recipients who were transplanted using this protocol with that of 68 matched ABO-compatible patients. In addition, we analyzed efficacy and cost of nonantigen-specific as compared to blood group antigen-specific IA. RESULTS: Before desensitization, the median isoagglutinin titer of 34 ABO-incompatible patients was 1:64 (Coombs technique). Patients received a median of 7 preoperative IA treatments. Twenty-four patients had a median of 2 additional plasmapheresis treatments to reach the preoperative target isoagglutinin titer of 1:8 or less. After a median postoperative follow-up of 22 months, overall graft survival in the ABO-incompatible group was not significantly different from that in ABO-compatible patients (log-rank P = 0.20), whereas patient survival tended to be lower (log-rank P = 0.05). The incidence of rejection episodes was 15% in both groups. The ABO-incompatible kidney recipients had a higher incidence of BK virus replication (P = 0.04) and nephropathy (P = 0.01) and showed more often colonization with multidrug resistant bacteria (P = 0.02). In comparison to blood group antigen-specific IA, nonantigen-specific IA showed equal efficacy but was associated with reduction in cost. CONCLUSIONS: Clinical outcomes of ABO-incompatible patients desensitized with a nonantigen-specific IA device and rituximab do not differ from that of matched ABO-compatible patients although a trend toward reduced patient survival was noted. Special attention must be paid to the higher incidence of BK virus infection in recipients of ABO-incompatible grafts.

Increased peritoneal damage in glyoxalase 1 knock-down mice treated with peritoneal dialysis.

BACKGROUND: Peritoneal dialysis (PD) is limited by peritoneal fibrosis and ultrafiltration failure. This is in part caused by the high concentration of glucose degradation products (GDPs) present in PD fluids (PDF) as a consequence of heat sterilization. Existing research in long-term PD has mainly dealt with the toxicity induced by GDPs and the development of therapeutic strategies to reduce the cellular burden of GDPs. Currently, there are few data regarding the potential role of detoxification systems of GDP in PD. In this study, the role of glyoxalase 1 (Glo1), the major detoxification pathway for dicarbonyl-derived GD such as methylglyoxal (MG) and glyoxal (Gx), was investigated in vivo using heterozygous knock-down mice for Glo1 (Glo1(-/+)). METHODS: Wild-type (WT) and Glo1(-/+) mice were repeatedly treated with PDF containing low and high amounts of GDP, particularly with respect to the content of dicarbonyls. After 12 weeks of treatment with PDF, peritoneal damage and function were evaluated. RESULTS: Glo1(-/+) mice treated with PDF showed increased formation of advanced glycation endproduct (AGE) when compared with WT mice, particularly the Gx-derived AGE, carboxymethyl-lysine. This was associated with increased inflammation, neovascularization, increased peritoneal fibrosis and impaired peritoneal function. CONCLUSIONS: This study suggests a pivotal and underestimated role for Glo1 as a detoxifying enzyme in GDP-associated peritoneal toxicity in PD. The indirect and direct modulation of Glo1 may therefore offer a new therapeutic option in prevention of GDP-induced peritoneal damage in PD.

Generation of suppressive blood cells for control of allograft rejection.

Our previous studies in rats showed that incubation of monocytic dendritic cells (DCs) with the chemotherapeutic drug mitomycin C (MMC) renders the cells immunosuppressive. Donor-derived MMC-DCs injected into the recipient prior to transplantation prolonged heart allograft survival. Although the generation of DCs is labour-intensive and time-consuming, peripheral blood mononuclear cells (PBMCs) can be easily harvested. In the present study, we analyse under which conditions DCs can be replaced by PBMCs and examine their mode of action. When injected into rats, MMC-incubated donor PBMCs (MICs) strongly prolonged heart allograft survival. Removal of monocytes from PBMCs completely abrogated their suppressive effect, indicating that monocytes are the active cell population. Suppression of rejection was donor-specific. The injected MICs migrated into peripheral lymphoid organs and led to an increased number of regulatory T-cells (Tregs) expressing cluster of differentiation (CD) markers CD4 and CD25 and forkhead box protein 3 (FoxP3). Tolerance could be transferred to syngeneic recipients with blood or spleen cells. Depletion of Tregs from tolerogenic cells abrogated their suppressive effect, arguing for mediation of immunosuppression by CD4⁺CD25⁺FoxP3⁺ Tregs. Donor-derived MICs also prolonged kidney allograft survival in pigs. MICs generated from donor monocytes were applied for the first time in humans in a patient suffering from therapy-resistant rejection of a haploidentical stem cell transplant. We describe, in the present paper, a simple method for in vitro generation of suppressor blood cells for potential use in clinical organ transplantation. Although the case report does not allow us to draw any conclusion about their therapeutic effectiveness, it shows that MICs can be easily generated and applied in humans.

Sulforaphane protects hearts from early injury after experimental transplantation.

BACKGROUND: Oxidative stress is a major factor in the development of ischemia reperfusion injury (IRI) after heart transplantation. The isothiocyanate found in broccoli - sulforaphane (SFN) - is an indirect antioxidant that acts by inducing Nrf2-dependent Phase 2 enzymes, such as NAD(P)H-quinone oxidoreductase 1, glutathione reductase, glutathione peroxidase, and the cardioprotective enzymes, haemoxygenase-1 (HO-1) and thioredoxin (Trx-1), participate in adaptive and protective responses to oxidative stress and various inflammatory stimuli. The aim of this study was to ameliorate IRI following heart transplants by recipient preconditioning. MATERIAL AND METHODS: Male Lewis rats were randomly divided into groups of n=10 animals each for heart donation. Heart grafts underwent 18 hours of cold storage in histidine-tryptophanketoglutarate (HTK) solution. Preconditioning of recipients with sulforaphane was performed 24 hours before transplantation. RESULTS: SFN significantly decreased serum levels of TnT, CK, CK-MB, LDH, AST, and ALT, which correlated with better graft function scores and improved survival prognosis. Pretreated recipients showed significantly decreased expression of iNOS, caspase 3, and HIF-1a. CONCLUSIONS: Our results indicate that recipient preconditioning with SFN protects the heart from IRI after experimental transplantation.

Kidney transplantation across HLA and ABO antibody barriers.

PURPOSE OF REVIEW: A significant number of kidney transplantations in industrialized countries is currently performed over human leukocyte antigen (HLA) or ABO antibody barriers after living donation to encounter the increasing shortage of organs from deceased donors. Although patients with moderate titers of anti-A/B antibodies may easily be desensitized with no negative impact on allograft survival, recipients with high titers and HLA sensitized patients demonstrate a substantial risk for antibody-mediated rejection, limiting long-term outcomes. RECENT FINDINGS: The use of powerful desensitization strategies including plasmapheresis and immunoadsorption, extended therapeutic options such as the application of the recently introduced complement inhibitors, and refined antibody detection techniques may further facilitate transplantations, especially in the HLA-sensitized kidney transplant recipient. On the contrary, special strategies such as the Eurotransplant Acceptable Mismatch Program or kidney paired exchange help improving long-term outcomes in these difficult to transplant patients by circumventing the HLA (or ABO) antibody barrier. SUMMARY: As compared with waiting for a compatible deceased donor organ, HLA and ABO incompatible transplantations performed in experienced centers have become a reasonable alternative for end-stage kidney disease patients with an incompatible live donor. Whenever possible, however, the transplantation should be performed between ABO compatible donor-recipient pairs in the absence of positive crossmatch results.

Cellular infiltrates and NFκB subunit c-Rel signaling in kidney allografts of patients with clinical operational tolerance.

BACKGROUND: Nuclear factor kappa B (NFκB) plays a potential role in tolerance by orchestrating onset and resolution of inflammation and regulatory T cell differentiation through subunit c-Rel. We characterized cellular infiltrates and expression of NFκB1, c-Rel and its upstream regulators phosphatidylinositol 3-kinase/RAC-alpha serine/threonine kinase, in allograft biopsies from patients with spontaneous clinical operational tolerance (COT). METHODS: Paraffin-fixed kidney allograft biopsies from 40 patients with COT (n=4), interstitial rejection (IR; n=12), borderline changes (BC; n=12), and long-term allograft function without rejection (NR; n=12) were used in the study. Cellular infiltrates and immunohistochemical expression of key proteins of the NFκB pathway were evaluated in the cortical tubulointerstitium and in cellular infiltrates using digital image analysis software. Results were given as mean±SEM. RESULTS: Biopsies from patients with COT exhibited a comparable amount of cellular infiltrate to IR, BC, and NR (COT, 191±81; IR, 291±62; BC, 178±45; and NR, 210±42 cells/mm) but a significantly higher proportion of forkhead box P3-positive cells (COT, 11%±1.7%; IR, 3.5%±0.70%; BC, 3.4%±0.57%; and NR, 3.7%±0.78% of infiltrating cells; P=0.02). c-Rel expression in cellular infiltrates was significantly elevated in IR, BC, and NR when analyzing the number of positive cells per mm (P=0.02) and positive cells per infiltrating cells (P=0.04). In contrast, tubular PI3K and c-Rel expression were significantly higher in IR and BC but not in NR compared with COT (P=0.03 and P=0.006, respectively). With RAC-alpha serine-threonine kinase, similar tendencies were observed (P=0.2). CONCLUSIONS: Allografts from COT patients show significant cellular infiltrates but a distinct expression of proteins involved in the NFκB pathway and a higher proportion of forkhead box P3-positive cells.

Effect of paricalcitol and calcitriol on aortic wall remodeling in uninephrectomized ApoE knockout mice.

Despite an only minor reduction in the glomerular filtration rate, uninephrectomy (UNX) markedly accelerates the rate of growth of atherosclerotic plaques in ApoE-/- mice. It has been suggested that vitamin D receptor (VDR) activation exerts an antiproliferative effect on vascular smooth muscle cells, but the side effects may limit its use. To assess a potentially different spectrum of actions, we compared the effects of paricalcitol and calcitriol on remodeling and calcification of the aortic wall in sham-operated and UNX ApoE-/- mice on a diet with normal cholesterol content. Sham-operated and UNX mice were randomly allotted to treatment with solvent, calcitriol (0.03 µg/kg) or paricalcitol (0.1 µg/kg) 5 times/wk intraperitoneally for 10 wk. Semithin (0.6 µm) sections of the aorta were analyzed by 1) morphometry, 2) immunohistochemistry, and 3) Western blotting of key proteins involved in vascular calcification and growth. Compared with sham-operated animals (5.6 ± 0.24), the wall-to-lumen ratio (x100) of the aorta was significantly higher in solvent- and calcitriol-treated UNX animals (6.64 ± 0.27 and 7.17 ± 0.81, respectively, P < 0.05), but not in paricalcitol-treated UNX (6.1 5 ± 0.32). Similar differences were seen with respect to maximal plaque height. Expression of transforming growth factor (TGF)-ß1 in aortic intima/plaque was also significantly higher in UNX solvent and UNX calcitriol compared with sham-operated and UNX paricalcitol animals. Treatment with both paricalcitol and calcitriol caused significant elevation of VDR expression in the aorta. While at the dose employed paricalcitol significantly reduced TGF-ß expression in plaques, calcitriol in contrast caused significant vascular calcification and elevated expression of related proteins (BMP2, RANKL, and Runx2).

In renal transplants with delayed graft function chemokines and chemokine receptor expression predict long-term allograft function.

BACKGROUND: Chronic loss of renal allograft function is associated with interstitial fibrosis and tubular atrophy (IF/TA). Independent of the underlying reason, one initial step in the development of fibrosis is chemokine-driven invasion of leukocytes from the blood vessels into the allograft. We studied the role of chemokines in kidney allografts with delayed graft function and the subsequent long-term outcome of renal function and fibrosis. METHODS: We examined repetitive biopsies of 30 patients without signs of acute rejection but with initially delayed graft function for IF/TA. In addition, we examined the expression of chemokine receptor (CCR)-1 and CCR2 on invaded leukocytes and macrophages and the corresponding ligands regulated upon activation, normal t-cell expressed, and secreted (RANTES) and monocyte chemotactic protein-1 on residential kidney cells. RESULTS: The initial expression of CCR1 positive invading cells and RANTES in glomerular cells correlated with the allograft function 12 months after transplantation and at last follow-up. The expression was independent of donor characteristics such as age, gender, infectious state, cause of death, or use of vasopressive agents. Furthermore, it did not correlate with the duration of cold ischemia time. Among the patients with the most progressive loss of allograft function follow-up biopsy specimen did not reveal any signs of rejection but showed increased CCR1 and RANTES expression in the interstitium suggesting ongoing inflammation and fibrosis. CONCLUSION: An early expression of RANTES in renal allografts with delayed graft function with consecutive invasion of CCR1 positive cells seems to promote ongoing IF/TA and to worse renal allograft outcome.

Does borderline kidney allograft rejection always require treatment?

BACKGROUND: Borderline rejection (Bord-R) is a frequent diagnosis in renal transplantation, and there is increasing evidence that regulatory T lymphocytes are involved in its pathogenesis. Current histopathologic practice does not differentiate between graft-protecting and -damaging T lymphocytes, and patients with Bord-R routinely receive rejection treatment. We analyzed Treg-associated forkhead box P3 (Foxp3) gene expression in Bord-R and more severe forms of acute rejection episodes (ARE). METHODS: Foxp3 transcripts were measured in 520 serial peripheral blood samples from 177 kidney graft recipients obtained during the first 20 days posttransplantation. RESULTS: The highest Foxp3 transcripts were observed in patients with Bord-R or without rejection and the lowest in patients with ARE. Patients with Bord-R on posttransplant days 5 to 7 showed an increased Foxp3 transcript level of 156%, which increased to 302% by posttransplant days 14 to 16. In contrast, patients with ARE demonstrated significantly lower Foxp3 gene expression than that observed in Bord-R, nonrejectors, or acute tubular necrosis patients (P=0.001, P<0.001, and P=0.005, respectively, on days 11-13). Acute tubular necrosis patients demonstrated intermediately high Foxp3 gene expression. CONCLUSIONS: Our data indicate that increased Treg activity in peripheral blood is a frequent feature of Bord-R. This finding questions the appropriateness of rejection treatment in all patients with the histopathologic diagnosis "Bord-R".

Clinical profile, predictors of mortality, and treatment of patients after myocardial infarction, in an academic medical center hospital.

OBJECTIVE: To evaluate clinical profiles, predictors of 30-day mortality, and the adherence to international recommendations for the treatment of myocardial infarction in an academic medical center hospital. METHODS: We retrospectively studied 172 patients with acute myocardial infarction, admitted in the intensive care unit from January 1992 to December 1997. RESULTS: Most patients were male (68%), white (97%), and over 60 years old (59%). The main risk factor for coronary atherosclerotic disease was systemic blood hypertension (63%). Among all the variables studied, reperfusion therapy, smoking, hypertension, cardiogenic shock, and age were the predictors of 30-day mortality. Most commonly used medications were: acetylsalicylic acid (71%), nitrates (61%), diuretics (51%), angiotensin-converting enzyme inhibitors (46%), thrombolytic therapy (39%), and beta-blockers (35%). CONCLUSION: The absence of reperfusion therapy, smoking status, hypertension, cardiogenic shock, and advanced age are predictors of 30-day mortality in patients with acute myocardial infarction. In addition, some medications that are undoubtedly beneficial have been under-used after acute myocardial infarction.

Perfil clínico, preditores de mortalidade e tratamento de pacientes após infarto agudo do miocárdio, em Hospital Terciário Universitário / Clinical profile, predictors of mortality, and treatment of patients after myocardial infarction, in an Academic Medical Center Hospital

OBJECTIVE: To evaluate clinical profiles, predictors of 30-day mortality, and the adherence to international recommendations for the treatment of myocardial infarction in an academic medical center hospital. METHODS: We retrospectively studied 172 patients with acute myocardial infarction, admitted in the intensive care unit from January 1992 to December 1997. RESULTS: Most patients were male (68 percent), white (97 percent), and over 60 years old (59 percent). The main risk factor for coronary atherosclerotic disease was systemic blood hypertension (63 percent). Among all the variables studied, reperfusion therapy, smoking, hypertension, cardiogenic shock, and age were the predictors of 30-day mortality. Most commonly used medications were: acetylsalicylic acid (71 percent), nitrates (61 percent), diuretics (51 percent), angiotensin-converting enzyme inhibitors (46 percent), thrombolytic therapy (39 percent), and beta-blockers (35 percent). CONCLUSION: The absence of reperfusion therapy, smoking status, hypertension, cardiogenic shock, and advanced age are predictors of 30-day mortality in patients with acute myocardial infarction. In addition, some medications that are undoubtedly beneficial have been under-used after acute myocardial infarction