José Gerardo Oliveira

Compared to mycophenolate mofetil, rapamycin induces significant changes on growth factors and growth factor receptors in the early days post-kidney transplantation.

BACKGROUND The new immunosuppressive drug Rapamycin (Rapa) is endowed with a mechanism of action that is distinct from that of calcineurin inhibitors. It has been claimed that Rapa does not significantly modulate either the cytokine expression or the transcription of several growth factors in mitogen-activated T cells. Previously, we reported that fine-needle aspiration biopsy (FNAB) sample cultures synthesize a large array of cytokines, and some of them may be powerful predictors of acute rejection in renal transplants. We hypothesized that Rapa may induce significant changes on cytokine production by FNAB sample cultures and on serum cytokine receptors when compared to other immunosuppressive drugs. METHODS Kidney transplants treated with CsA-Rapa-Pred (Rapa group) were compared with transplants treated with CsA-mycophenolate mofetil-Pred (MMF group). They were studied on day 7 posttransplantation, and they remained rejection free for at least the first 6 months. FNAB samples were cultured and the supernatants were collected at 48 hr of incubation and analyzed by ELISA for interleukin 1 receptor antagonist (IL-1ra), IL-2, IL-6, IL-10, IL-18, monocyte chemotactic protein 1 (MCP-1), soluble tumor necrosis factor I, and transforming growth factor (TGF)-beta(1). The soluble receptors for IL-1, IL-2, IL-6, and tumor necrosis factor alpha, together with IL-2 and IL-18 were also measured in serum. RESULTS Significant differences were observed when comparing Rapa with the MMF group. IL-18 and TGF-beta(1) synthesis were up-regulated, whereas IL-6 and MCP-1 were down-regulated in FNAB sample cultures. The Rapa group showed a significant down-regulation of each cytokine receptor and of IL-2 in serum. CONCLUSIONS Rapa was associated with a decreased synthesis of primarily monocyte-derived cytokines and enhanced production of TGF-beta(1), which in an appropriate cytokine milieu may promote allograft tolerance. The down-regulation of cytokine receptors and IL-2 may be associated with a depressed immune response towards the kidney allograft.

Analysis of fine-needle aspiration biopsies by flow cytometry in kidney transplant patients

BACKGROUND Peripheral blood lymphocyte (PBL) analysis by flow cytometry has been inconsistently reported as an adjunctive method for diagnosing acute kidney transplant rejection. However, there is good evidence that lymphocytes infiltrating renal grafts differ from those found at the peripheral level. We hypothesized that the study of aspiration biopsy samples in conjunction with PBL by flow cytometry would enable us to diagnose acute rejection crisis reliably. METHODS Lymphocytes from PBL and aspiration biopsies of kidney transplant patients were analyzed. Fifty-one stable patients, rejection-free for the first 6 months, were studied on day 7 and day 30 after transplantation and were compared with 32 patients with 40 acute rejection episodes. RESULTS Significant differences were observed for several lymphocyte subpopulations on aspiration biopsy samples comparing stable patients with rejection patients. In contrast, PBL analysis was not helpful in differentiating the two groups of patients. By combining the expression of several activation markers inside the graft with CD3DR and CD3CD25 aspiration biopsy to peripheral blood ratios, we obtained very good values for sensitivity and specificity-83.9% and 90.5%, respectively. The positive predictive value for rejection among dysfunctional grafts reached 85.8%. CONCLUSIONS Flow cytometry study of aspiration biopsy samples of kidney transplant patients is a reliable and powerful method to diagnose acute rejection episodes, although it is needed to consider several lymphocyte phenotypes; cytofluorometric analysis of PBL is important because it provides graft-infiltrating cell to peripheral blood lymphocyte ratios. This safe and rapid test may significantly improve the management of kidney transplant patients.

The synthesis by fine-needle aspiration biopsy cultures of IL-7, IL-16 and IL-18 is significantly associated with acute rejection in kidney transplants.

Background: T-cell activation, the key event in the development of acute allograft rejection, depends on co-stimulatory signals delivered by antigen-presenting cells (APC). APC-derived cytokines may provide co-stimulation and modulate alloimmune reaction. We have studied cytokine synthesis by fine-needle aspiration biopsy (FNAB) culture and we found significant differences for interleukin (IL)-2, IL-6, IL-10, M-CSF and IL-1ra on comparing acute rejection versus stable kidney transplant patients. We report our findings on FNAB cultures synthesis of IL-7, IL-15, IL-16, IL-17, IL-18 and RANTES (regulated upon activation, normal T-cell expressed and secreted), all potential modulators of anti-graft reaction. Patients and Methods: Kidney transplants (KTX) treated with CsA-AZA-Pred from the beginning, were divided into four groups. Group I: day 7 post-KTX, stable; II: day 7 post-KTX, 6.5 ± 5.5 days before acute rejection; III: first day of acute rejection; IV: day 14 post-KTX, stable. Patients from I and IV remained rejection-free for the first 6 months, at least. All rejection episodes were confirmed by classical core renal biopsy. FNAB samples were cultured according to our published methodology and culture supernatants were collected at 48 h and analysed by ELISA for IL-7, IL-15, IL-16, IL-17, IL-18 and RANTES. Results: Group III synthesized significantly higher amounts of IL-7, IL-16 and IL-18 than stable patients (groups I and IV). RANTES production did not show significant differences among the four groups. We did not find any trace of IL-15. Conclusions: IL-18 may play the activation role that has been attributed to IL-12 which previously, we did not find to correlate significantly with acute rejection in KTX. IL-16 seems to play an activation role rather than an inhibition of anti-graft reaction. We confirm that RANTES is not significantly associated with acute rejection in KTX.

Type of vascular access and location in online hemodiafiltration and its association with patient's perception of health-related quality of life.

Purpose The aim of this work is to evaluate the patient-reported health-related quality of life (HRQOL), according to the type and location of vascular access used for dialysis procedure. Methods In this transversal study, 322 end-stage renal disease (ESRD) patients under online hemodiafiltration (OL-HDF, 59.63% males; 64.9±14.3 years) were enrolled. Arteriovenous fistula (AVF) was used by 252 patients (78.3%), whereas 70 patients (21.7%) had a central venous catheter (CVC). Besides AVF location, data on comorbidities, hematological data, iron status, dialysis adequacy, nutritional and inflammatory markers were collected. Moreover, the patients’ reported HRQOL score, using the Kidney Disease Quality of Life-Short Form, was evaluated. Results ESRD patients using CVC as vascular access presented a decrease in four SF-36 domain scores, namely physical functioning, emotional well-being, role-emotional and energy/fatigue when compared with those using AVF as vascular access. Additionally, these patients also showed significant differences in ESRD target areas, namely decline in cognitive function and quality of social interaction domains. When comparing the variables according to the localization of the AVF, significant differences were found in three SF-36 domain scores, namely physical functioning, pain and general health. Moreover, we also found significant differences in ESRD target areas, namely symptoms/problem list, effects of kidney disease and quality of social interaction domains. Conclusions Our results showed that ESRD patients under OL-HDF using AVF as vascular access had higher HRQOL scores in several domains when compared with those using CVC. Additionally, we also found that dialysis patients using AVF in the left forearm presented with higher HRQOL scores.

sTNFRI and sTNFRII synthesis by fine-needle aspiration biopsy sample cultures is significantly associated with acute rejection in kidney transplantation.

Background. Previously we reported that cultures of fine-needle aspiration biopsy (FNAB) samples synthesize different cytokine pattern depending on the alloimmune response towards the kidney graft. However, we failed to find a clear picture for growth factors implicated in early T-cell activation (interferon-&ggr;, interleukin [IL]-4, IL-12), although we observed that interleukin-1 receptor antagonist (IL-1ra) was associated with absence of acute rejection. We have now studied tumor necrosis factor-&agr; (TNF-&agr;) and its two soluble receptors, sTNFRI and sTNFRII, IL-1&bgr; and soluble IL-1 receptor II (sIL-1RII), and leukemia inhibitory factor (LIF), all potential modulators of T-cell activation. Methods. Sixty-six cadaver kidney transplants (KTX) were divided into four groups: group 1, day 7 after KTX, stable (n=30); group 2, day 7 after KTX, 8±4.5 days before acute rejection (n=12); group 3, first day of acute rejection (n=17); and group 4, day 14 after KTX, stable (n=32). Patients from groups 1 and 4 remained rejection-free for the first 6 months. All rejection episodes were confirmed by core renal biopsy. FNAB samples were cultured according to our published methodology, and culture supernatants were collected at 48 hr and analyzed by ELISA for IL-1&bgr;, sIL-1RII, TNF-&agr;, sTNFRI, sTNFRII, and LIF. Serum levels for sIL-1RII, sTNFRI, and sTNFRII were also measured. Results. FNAB cultures from groups 1 and 4 synthesized significantly lower amounts of sTNFRI and sTNFRII than those from either groups 2 or 3. Both sTNFRI and sTNFRII reached high positive and negative predictive values for acute rejection. IL-1&bgr; and sIL-1RII were synthesized by all groups but without differences. No trace of LIF and TNF-&agr; was found. sTNFRII was significantly higher in serum from group 3. Conclusions. Both TNF receptors were positively associated with acute rejection and were good predictors of impending acute rejection. The ratio of sIL-1RII over IL-1 (together with IL-1ra that we previously measured in FNAB cultures) suggests that IL-1 actions may be inhibited with current immunosuppression early after transplantation.

Relationship between everolimus blood concentration assessed using the Innofluor Certican Fluorescence Polarization Immunoassay and the Architect i System sirolimus chemiluminescent microparticle immunoassay.

Everolimus, an immunosuppressive macrolide derivative of sirolimus, has a narrow therapeutic index and variable bioavailability. Assessment of blood concentration of everolimus is necessary to improve immunosuppressive efficacy without increasing potential adverse effects. Recently, Seradyn, Inc (Indianapolis, Indiana) introduced a fluorescence polarization immunoassay (Innofluor Certican Fluorescent Polarization Immunoassay [FPIA]) for quantitation of everolimus blood concentration. This immunoassay has concentration-dependent cross-reactivity with sirolimus, which must be considered in patients recently treated with that drug. In this short-term study, treatment in 53 renal transplant recipients was converted from a sirolimus-based regimen to an everolimus-based regimen. Patients were followed up for 3 months. We investigated whether cross-reactivity with everolimus also occurred with the Abbott Laboratories (Abbott Park, Illinois) Architect i System, a sirolimus chemiluminescent microparticle immunoassay (CMIA) used to quantify sirolimus blood concentration. Quantification of everolimus blood concentration using both the CMIA and the FPIA demonstrated a linear regression: CMIA = 0.73 FPIA +/- 0.77 (r(2) = 0.80; P < .001). A high degree of correlation between the CMIA and FPIA methods (r = 0.90) was confirmed using the Bland-Altman test. We conclude that the Abbott Architect i System sirolimus CMIA should be considered an alternative method for everolimus drug monitoring. The cross-reactions of both the FPIA and CMIA techniques with both sirolimus and everolimus must be considered when converting therapy from one drug to the other. In these conditions, use of an equivalent dosage is of particular importance.

Humoral immune response after kidney transplantation is enhanced by acute rejection and urological obstruction and is down-regulated by mycophenolate mofetil treatment

The anti‐allograft immune response may have a cellular and a humoral component. Lymphocytotoxic antibodies (Ab) and anti‐human leucocyte antigen (HLA) Ab present before kidney transplantation carry an enhanced risk of acute rejection. Current immunosuppressive drugs act predominantly upon the cellular immune pathway which may leave unopposed the humoral mechanisms of anti‐allograft response. We studied the production of lymphocytotoxic Ab and anti‐HLA Ab after kidney transplantation under different drug therapies. Two hundred and sixty‐four consecutive kidney transplant recipients treated with different immunosuppressive drugs, either stable and or with previous acute rejection or acute urologic obstruction, entered this study. Lymphocytotoxic Ab and anti‐HLA Ab were evaluated by complement‐dependent cytotoxicity and by ELISA. Ab donor‐specificity was determined by flow cytometry. Both lymphocytotoxic Ab and anti‐HLA Ab were significantly increased in acute rejection whatever the immunosuppressive regimen and almost significantly in urologic obstruction treated with azathioprine (AZA) groups. The presence of antidonor‐specific Ab was associated with a significantly higher rate of graft loss. Mycophenolate mofetil (MMF) therapy significantly down‐regulated Ab synthesis in all patients groups when compared with AZA. The development of humoral antidonor response post‐transplantation is associated with a dismal graft prognosis. This is the first report that acute urologic obstruction may be followed by unspecific lymphocytotoxic and anti‐HLA Ab synthesis, surmising that a protracted obstruction may promote renal fibrosis through antibody mediation. The significant down‐regulation of the humoral response by MMF when compared with AZA may herald a lower risk to mount a chronic rejection process.

Body composition assessed by impedance changes very early with declining renal graft function.

Background: Kidney transplant (Tx) restores renal filtration, although it does not achieve the function of two native kidneys, and with time it may variably involute back to chronic renal failure. We hypothesized that bioelectrical impedance analysis (BIA) might highlight differences for body compartments among Tx with different filtration rates, and we compared them with healthy controls. Methods: 38 Tx patients (25 males, 13 females) were studied at 75.9 ± 37.8 months postsurgery and divided into three groups: good creatinine clearance (CrCl, ml/min/1.73 m2; > 65.0), borderline (35.0 < CrCl < 60.0) and bad (CrCl < 35.0). BIA was assessed three times in a year. Total body water, extracellular water (ECW), intracellular water (ICW), Na:K exchange rate (Nae:Ke) and phase angle were studied. Healthy (n = 11) and hemodialysis (n = 11) groups were also studied. Results: BIA showed no differences between healthy controls and good Tx while both borderline and bad Tx presented a significantly higher ECW and lower ICW than either good Tx or normal controls. Only good CrCl was different from predialysis. Conclusions: A good kidney graft manages to restore and maintain normal body composition, even with potential disturbances brought about by steroids and cyclosporine. With mild renal dysfunction a change in body compartments was observed, moving towards the composition of that with chronic renal failure patients.

Salt sensitivity of blood pressure in patients with psoriasis on ciclosporin therapy

Background  Hypertension is one of the main side‐effects of long‐term therapy with ciclosporin. However, the influence of salt intake on the 24‐h mean blood pressure of patients with psoriasis treated with ciclosporin is not known.

Risk factors for mortality in end-stage kidney disease patients under online-hemodiafiltration: three-year follow-up study

Abstract The aim of this work was to evaluate the predictors of mortality in a group of end-stage kidney disease (ESRD) patients under dialysis, by performing a three-year follow-up study. From the 236 patients included in this study, 54 patients died during the three-year follow-up period. Our data showed that the risk of death was higher in patients presenting lower levels of mean cell hemoglobin concentration, transferrin, and albumin. Our study showed that poor nutritional status and an inflammatory-induced iron depleted erythropoiesis are important factors for mortality in these patients.