Long-term Glomerular Filtration Rate and Kidney Disease: Improving Global Outcomes Stage Stability After Conversion to Once-Daily Tacrolimus in Kidney Transplant Recipients.

Close monitoring of estimated glomerular filtration rate (eGFR) is important for early recognition of worsening renal function to prevent further deterioration. Safe conversion from twice-daily tacrolimus (TD-Tac) to once-daily tacrolimus (OD-Tac) has been reported, but the effects on eGFR are contrasting. The aim of our study is to evaluate long-term stability of eGFR after 1:1 conversion from TD-Tac to OD-Tac and the effects on serum cytokine blood levels. Forty-six consecutive kidney transplant recipients treated with TD-Tac 3 to 5 years post-transplant, with stable renal function, were enrolled in the study (2009-2011). Clinical and biochemical parameters were evaluated for 12 months before conversion up to 6 years after conversion. The patients served as their own controls. A panel of cytokines was evaluated repeatedly during the first year after conversion. Mean values of eGFR were not different long-term after conversion (P = .11) compared with baseline, and the majority of patients remained stable on Kidney Disease: Improving Global Outcomes stage during the study period; eGFR was stable in 30.0% after 5 years, decreased > 1 mL/min/1.73 m2/y in 13.3%, and improved > 1 mL/min/1.73 m2/y in 56.7%. Cytokine levels and C-reactive protein did not show any significant deterioration. Metabolic parameters were stable during the 6 years of follow-up. OD-Tac therapy can preserve an effective immunosuppressive state together with a safe profile of eGFR.

Automated Intelligent Microscopy for the Recognition of Decoy Cells in Urine Samples of Kidney Transplant Patients.

BACKGROUND: BK virus (BKV)-associated nephropathy is definitely involved in allograft failure after kidney transplant. Thus, the need for an early control of viral reactivation in immunocompromised patients is well established. Determination of urinary release of decoy cells (DC) and BK viral load in plasma and urine by polymerase chain reaction (PCR) usually precedes renal biopsy. The aim of the study is to assess viral reactivation by BKV-DNA PCR and DC detection in urinary sediment using automated intelligent microscopy. METHODS: Seventy-eight kidney transplant patients were analyzed for the presence of plasma BKV-DNA by quantitative TaqMan real-time PCR. Additionally, automated intelligent microscopy was used for urine sediment analysis, allowing to count cells with decoy feature, confirmed by phase contrast microscopic review. RESULTS: Plasma BKV-DNA PCR was detected in 14 (17.9%) patients. DC were identified in 19 (24.3%) urine sediments by automated analyzers and confirmed by microscopic observation. Two patients were BKV-DNA-positive/DC-negative; conversely, 7 subjects were DC-positive/BKV-DNA-negative. CONCLUSIONS: Plasma quantification of BK viral load is currently the best noninvasive method for the detection of viral reactivation. Nevertheless, automated methods to screen for the presence of DC in urine could facilitate early BK virus replication diagnosis and patient follow-up by quantitative and visual results.

Role of biomarkers in adult sepsis and their application for a good laboratory practice: a pilot study.

This study measured Procalcitonin (PCT), Presepsin (PRE-S) and pro-Adrenomedullin (pro-ADM) in intensive care unit (ICU) patient’s blood to assess their contribution to accurate diagnosis of sepsis and potential predictive impact on prognosis. The final aim was to improve the use of infection biomarkers for optimizing the impact of laboratory medicine on clinical outcomes, focusing on the good management of resources designed to produce maximum effectiveness and efficiency. Sixty-four adult patients were studied during their hospitalization in ICU; blood samples were collected and categorized according to their clinical diagnosis and illness severity, and sepsis marker levels were measured on automated immunoassay platforms. PCT, PRE-S and pro-ADM infection markers were significantly lower in controls than in sepsis or septic shock groups. The area under the curve, by ROC curve analysis, was 0.945 for PCT, 0.756 for PRE-S and 0.741 for pro-ADM. Sepsis diagnostic accuracy was not improved by combining PCT, PRE-S and pro-ADM measures. Preliminary data demonstrated that, despite PRE-S and pro-ADM being able to differentiate between septic and non-septic patients with accuracy, PCT remains the most reliable marker available. The results obtained still do not allow us to consider a combination of markers, because it would merely increase laboratory costs without improving diagnostic performance. Furthermore, the results confirm a possible prognostic role of pro-ADM in septic states, but no correlation between biomarker levels and survival at 48 h was detected. Hence PCT, PRE-S, nor pro-ADM can be used to predict short-term prognosis.

Assessment of NGAL as an early biomarker of acute kidney injury in adult cardiac surgery patients.

Early and predictive acute kidney injury (AKI) markers may be decisive for the clinical outcome of heart surgery. Hence, this study set out to evaluate the biological variability of urinary neutrophil gelatinase-associated lipocalin (uNGAL) levels in adult cardiac surgery patients, to test their feasibility as a biomarker of early AKI in a routine laboratory setting. uNGAL levels were measured with an automated immunoassay in urine samples from patients undergoing cardiac surgery using cardiopulmonary bypass, at the time of admission (T0) and 4 hours (T1) and 24 hours (T2) after surgery. Patients without post-operative AKI did not show significant differences in urine NGAL levels after surgery. In contrast, patients developing AKI displayed a significant increase (P=0.011) in uNGAL levels compared to T0. This increase was detectable at an earlier time point (T1, 4 hours) with respect to serum creatinine (T2, 24 hours). Confirming its utility as a biomarker, at T1 the uNGAL levels were significantly higher in AKI patients than in non-AKI patients (P=0.021). A receiver operating characteristic curve analysis of the uNGAL assay gave a sensitivity of 55.3 (95percent confidence interval, 26.59-78.73), a specificity of 72.9 (95 percent CI, 55.88-86.21), and a cut-off value for AKI prediction of 55.2. These results support the notion that urinary NGAL is an earlier marker of AKI than serum creatinine. However, the cut-off value of the assay was too low to consider it as a positive or negative diagnostic marker in AKI patients with moderate degree of severity. Likewise, its sensitivity and specificity were not high enough for it to be considered better than the others currently in use.

Long-term interleukin-2 assessment after conversion from a twice-daily to once-daily tacrolimus regimen in stable kidney recipients.

Conversion to tacrolimus (Tac) to once daily (Tac-O) formulation is commonly followed by a 20% reduction in Tac trough levels in the first month. It is not associated with modifications of renal function but there is the issue of its effects on inflammatory cytokines and on subclinical rejection. The aim of our study was to evaluate long-term interleukins (IL)-2 profiles in stable renal transplant patients after Tac-O conversion. We enrolled 10 stable kidney transplant patients converted to Tac-O. Tac trough levels, serum creatinine concentrations, glomerular filtration rate using the Modification of Diet in Renal Disease formula, C-reactive protein, IL-2 levels, and clinical assessments were performed monthly for 6 months before and 12 months after conversion. Despite the significant reduction in Tac trough levels, we did not observe alterations suggestive of clinical or subclinical acute rejection.

Interleukin-2 profiles shortly after tacrolimus conversion from a twice-daily to once-daily regimen.

A number of studies have indicated that kidney recipients can be safely converted from the twice-daily formulation (Tac-T) to the same dose of a once-daily tacrolimus (TAC) regimen (Tac-O) based upon monitoring of renal function. Conversion from Tac-T to Tac-O is commonly followed by a reduction in Tac trough levels, estimated by some authors to be about 20%. These alterations seem to not be associated with a modification of graft function, but study of inflammatory cytokines would be useful. The aims of our study were to monitor Tac, C-reactive protein (CRP), and interleukin (IL)-2 levels as well as to evaluate renal function among stable renal transplant patients converted from a Tac-T to a Tac-O regimen. We enrolled 10 consecutive stable kidney transplanted patients. Tac trough levels, serum creatinine concentrations, glomerular filtration rates using the Modification of Diet in Renal Disease formula (MDRD), CRP, and clinical assessment were performed monthly for 6 months before and 3 months after the conversion. After conversion we observed a slight but not significant reduction in Tac trough level. Renal function evaluated by serum creatinine and MDRD as well as CRP were not significantly different after conversion. IL-2 levels remained stable after conversion. We identified a group of patients showing reduced Tac trough levels below the therapeutic range and a group with stable Tac levels. No significant differences were observed among the two groups before versus after the conversion. Our results did not show a modification of IL-2, CRP and renal function levels, at 3 months after conversion despite the lower Tac trough concentrations. The clinical meaning of Tac trough alterations is not clear. They might reflect inter- and intraindividual differences in the clearance of Tac as recently described. They did not seem to be associated with activation of an inflammatory pathway.

Tacrolimus trough levels and level-to-dose ratio in stable renal transplant patients converted to a once-daily regimen.

Numerous evidence has been reported to support a safe 1:1 conversion from the twice-daily tacrolimus (Tac-T) to the once-daily tacrolimus regimen (Tac-O), but frequently there is a reduction in drug trough levels, which has been estimated by some authors to be about 20%. The relationship between Tac-O dosage and trough levels after conversion is not clear. The tacrolimus trough levels-to-dose ratio has been applied to better define the wide variability in doses and blood levels of tacrolimus. The aim of this study was to evaluate tacrolimus trough levels, tacrolimus daily dosage, and tacrolimus level-to-dose ratio during 1 year pre-postconversion follow-up in 31 stable kidney transplant patients who had received Tac-T therapy for over 6 months with stable renal function. They were converted to the same dosage of Tac-O. Patients before and after conversion were their own controls. The trough levels of tacrolimus showed a slight albeit significant reduction after conversion, remaining in the therapeutic range. Nineteen percent underwent an adjustment in total daily dosage after conversion versus 39% before conversion with no significant difference. No significant differences were detected in the total daily dose administered either by tacrolimus level-to-dose ratio before or after conversion. Kidney transplant recipients under Tac-O therapy were safely maintained using the same therapeutic monitoring as when receiving Tac-T.

Improvement of graft function after conversion to once daily tacrolimus of stable kidney transplant patients.

BACKGROUND: Chronic renal dysfunction is present in about one quarter of kidney transplant patients at 1 year and in about 90% by 10 years. Nephrotoxicity caused by calcineurin inhibitors is among the most common factors. Elevated tacrolimus levels have been correlated with worse control of side effects including acute and/or chronic nephrotoxicity. The aim of this study was to observe the effects on graft function of conversion from the twice daily to the once daily extended release tacrolimus formulation in stable kidney transplant recipients within 5 years of grafting. METHODS: Thirty-one stable kidney transplant patients were converted at the same dosage (1 mg:1 mg). Patients served as their own controls based on results before versus after conversion. RESULTS: The trough levels of tacrolimus showed a slight albeit significant reduction after the conversion. Serum creatinine and glomerular filtration rate showed a significant improvement without an association with the tacrolimus trough levels. CONCLUSION: We suggest that the immunosuppression with once daily tacrolimus may be a good option for kidney transplant patients.

Once daily tacrolimus formulation: monitoring of plasma levels, graft function, and cardiovascular risk factors.

BACKGROUND: Advagraf, an extended release formulation of tacrolimus, is administered once daily during the morning fast. Tacrolimus can be safely converted from the twice daily formulation (Prograf) to the same dose (1 mg:1 mg) of once daily dosing tacrolimus (m-Tac). The adverse effects of tacrolimus play important roles in posttransplant cardiovascular risk factors (CVR): hyperglycemia, posttransplant diabetes mellitus, dyslipidemia and hypertension. It has been suggested that avoiding high tacrolimus peak levels minimizes its diabetogenic effects leading to better glycemic control. The aim of our study was to observe the effects of conversion to m-Tac therapy on graft function and CVR among stable transplant kidney recipients. METHODS: We selected 2 groups of 20 patients with stable kidney transplantation, who had been treated with Prograf for >6 months with a triple regimen. Group 1 were converted to once daily tacrolimus at the same dose (1 mg:1 mg); whereas in group 2, the therapy was maintained as a twice daily regimen. Blood pressure, creatinine and glomerular filtration rate levels evaluated by the Modification of Diet in Renal Disease formula, as well as urea, total, high- and low-density lipoprotein remained stable between the 2 groups as well as inside group 1 before and after conversion. RESULTS: After conversion, glycemia and triglyceride values showed significant reductions in group 1 and between the 2 groups. These results were significant, as they may be associated with better long-term graft and patient survivals.

Preliminary evaluation of the new TACR flex method versus MEIA method in the therapeutic monitoring of tacrolimus in organ transplantation.

Tacrolimus (FK506) is an effective macrolide immunosuppressant widely used to prevent organ rejection following transplantation. Monitoring blood levels of tacrolimus is essential to assess organ rejection versus toxicity, because of the narrow therapeutic range and pharmacokinetic variability. The increased request for therapeutic drug monitoring is an interesting challenge for clinical laboratories. The automated immunoassay methods provide correct results and a turnaround time considerably reduced compared to HPLC and HPLC-MS which remain the gold standard for accuracy and economical advantages. A new immunoassay method, TACR Flex Dimension, is a commercially available, automated pretreatment test. The purpose of this study was to compare two analytical methods: the MEIA II tacrolimus immunoassay using the IMx analyzer and the new TACR Flex tacrolimus immunoassay on the Dimension system. Tacrolimus results obtained using the two methods were compared using European control and 93 whole blood samples from kidney and liver transplant patients. The tacrolimus concentrations measured by Flex Dimension for all samples were higher (0.7 to 16.1 ng/mL) than results obtained with MEIA (0.2 to 13.4 ng/mL), a mean difference expressed in percentage of 31.7%, and a correlation coefficient of 0.85. The data obtained by both methods using three European controls showed similar concentrations. The TACR Flex Dimension method provided a higher automation level and therefore a lower incidence of preanalytical errors and a lower turnaround time.

Effector Th-1 cells with cytotoxic function in the intestinal lamina propria of patients with Crohn's disease.

A large body of evidence points to a pivotal relationship between Th-1 cells and mucosal inflammation in Crohn's disease (CD). The aim of the present study was to assess whether CD is associated with specific functional activity of lamina propria T lymphocytes (LPT), particularly purified CD4, such as cytotoxic activity and specific cytokine-secreted profile. The results showed that CD4 LPT in patients displayed a chronically activated memory-like surface phenotype and, when compared to controls, had a significantly enhanced antibody-redirected cytotoxicity. Interestingly, the ratio of perforin expression in CD4 LPT was higher compared to controls, and a redirected lysis of human RBC mediated by a CD4 subset of intestinal lamina propria was evident, suggesting a cytolytic pore-forming mechanism. Moreover, a unique Th-1 cytokine profile pattern in the CD4 cells from CD was defined. These effector cells produced 12 times more IFN-gamma, two times more TNF-alpha, and three times less IL-4 than controls. In contrast, no increase in IL-2 was detected, while IL-5 was undetectable. Our studies suggest that these preexisting in vivo activated CD4 LPT may play an important role in the inflammatory process in CD, thus directly contributing to the intestinal lesions.

Homing of CD4+CD56+ T lymphocytes into kidney allografts during tubular necrosis or rejection.

The association between acute rejection, acute tubular necrosis, good function and relative infiltration of CD56 subsets of both CD8+ and CD4+ T cells was examined on 67 samples of graft infiltrating cells (GIC) and corresponding peripheral blood lymphocytes (PBL) obtained from renal allograft recipients. Quantification of cell subset profiles was determined by two-color flow cytometry. While a high proportion of CD4+CD56+ GIC was detected when both renal dysfunction and graft cytopathology (acute tubular necrosis or acute rejection) were present, this cell subset was undetectable in peripheral blood. In contrast the CD8+CD56+ T-cell subset was not discriminatory. The presence of CD4+CD56+ cells among freshly-isolated lymphocytes from renal allografts supports the idea that the local environment is involved in the selection of this subset, thus participating in the amplification of the immune-response. In addition, a homing of this T-cell subset into the transplanted organ may constitute an early sign of graft immunopathology.

Efficacy of cyclosporine to reduce steroids in patients with idiopathic pulmonary fibrosis before lung transplantation.

Lung transplantation today is considered an effective option for patients with severe idiopathic pulmonary fibrosis. The standard medical treatment for this disease consists of high-dose steroids alone or combined with other immunosuppressive drugs. Unfortunately, pretransplantation administration of steroids may jeopardize the healing of the airway anastomosis and cause other complications; therefore it is considered a relative contraindication to lung transplantation. For this reason we try to reduce the dose of prednisone to 15 to 20 mg/day or less before the transplantation, but this creates many difficulties and is sometimes impossible in severely ill patients. Therefore we used cyclosporine (4 to 7 mg/kg/day) in 10 patients who were receiving high-dose prednisone (> or = 50 mg/day) therapy, but who were otherwise suitable candidates for lung transplantation. In seven cases prednisone could be tapered to 20 mg/day or less, allowing acceptance in our program. These patients had a CRP score of 60 or more before entering our trial and remained stable at this level after conversion to cyclosporine. The 6-minute walk test showed a mild improvement in five cases (71.5%). Three patients underwent single lung transplantation; two patients are on our waiting list after 3.5 and 4 months of treatment with cyclosporine and prednisone (10 mg/day), and two patients died while awaiting a suitable organ 6 and 7.5 months after starting cyclosporine therapy. Combined administration of cyclosporine and prednisone may extend the waiting time while receiving low-dose steroids and allow more patients with idiopathic pulmonary fibrosis to qualify for lung transplantation while reducing the risk of steroid-induced complications.