Paola Todeschini

VDR Expression on Circulating Endothelial Progenitor Cells in Dialysis Patients Is Modulated by 25(OH)D Serum Levels and Calcitriol Therapy

Background/Aims: Vitamin D deficiency is associated with endothelial dysfunction in uremic patients, possibly by the impairment in the number and function of endothelial progenitor cells (EPCs). In 89 hemodialysis patients, we investigated the factors associated with the number of circulating EPCs (CD34⁺/CD133⁺/KDR⁺ and CD34⁺/CD133^–/KDR⁺ cells), the presence of VDR and the determinants of VDR expression on EPCs, in particular in calcitriol therapy. Methods: EPC counts, percentages of VDR-positive EPCs and VDR expression were assessed by flow cytometry. Cells isolated from a subgroup of patients were cultured to analyze colony-forming units, specific markers expression and a capillary-like structure formation. Results/Conclusions: Our study demonstrates the presence of VDR on EPCs. In our dialysis patients, the parameters studied on both CD34⁺/CD133⁺/KDR⁺ and CD34⁺/CD133^–/KDR⁺ cells, in particular VDR expression, seem to be influenced by uremia-related factors, including anemia, inflammation, diabetes, 25(OH)D serum levels and calcitriol therapy.

Outcomes of Pregnancies after Kidney Transplantation: Lessons Learned from CKD. A Comparison of Transplanted, Nontransplanted Chronic Kidney Disease Patients and Low-Risk Pregnancies: A Multicenter Nationwide Analysis

Background Kidney transplantation (KT) may restore fertility in chronic kidney disease (CKD). The reasons why maternofetal outcomes are still inferior to the overall population are only partially known. Comparison with the CKD population may offer some useful insights for management and counselling. Aim of this study was to analyse the outcomes of pregnancy after KT, compared with a large population of nontransplanted CKD patients and with low-risk control pregnancies, observed in Italy the new millennium. Methods We selected 121 live-born singletons after KT (Italian study group of kidney in pregnancy, national coverage about 75%), 610 live-born singletons in CKD, and 1418 low-risk controls recruited in 2 large Italian Units in the same period (2000-2014). The following outcomes were considered: maternal and fetal death; malformations; preterm delivery; small for gestational age (SGA) baby; need for the neonatal intensive care unit; doubling of serum creatinine or increase in CKD stage. Data were analyzed according to kidney diseases, renal function (staging according to CKD-epidemiology collaboration), hypertension, maternal age, parity, ethnicity. Results Maternofetal outcomes are less favourable in CKD and KT as compared with the low-risk population. CKD stage and hypertension are important determinants of results. Kidney transplantation patients with estimated glomerular filtration rate greater than 90 have worse outcomes compared with CKD stage 1 patients; the differences level off when only CKD patients affected by glomerulonephritis or systemic diseases (“progressive CKD”) are compared with KT. In the multivariate analysis, risk for preterm and early-preterm delivery was linked to CKD stage (2-5 vs 1: relative risk 3.42 and 3.78) and hypertension (RR 3.68 and 3.16) while no difference was associated with being a KT or a CKD patient. Conclusions The maternofetal outcomes in patients with kidney transplantation are comparable with those of nontransplanted CKD patients with similar levels of kidney function impairment and progressive and/or immunologic kidney disease.

Neutrophil Gelatinase-Associated Lipocalin Increases HLA-G+/FoxP3+ T-Regulatory Cell Population in an In Vitro Model of PBMC

Background Neutrophil gelatinase-associated lipocalin (NGAL) is emerging as a mediator of various biological and pathological states. However, the specific biological role of this molecule remains unclear, as it serves as a biomarker for many conditions. The high sensitivity of NGAL as a biomarker coupled with relatively low specificity may hide important biological roles. Data point toward an acute compensatory, protective role for NGAL in response to adverse cellular stresses, including inflammatory and oxidative stress. The aim of this study was to understand whether NGAL modulates the T-cell response through regulation of the human leukocyte antigen G (HLA-G) complex, which is a mediator of tolerance. Methodology/Principal Findings Peripheral blood mononuclear cells (PBMCs) were obtained from eight healthy donors and isolated by centrifugation on a Ficoll gradient. All donors gave informed consent. PBMCs were treated with four different concentrations of NGAL (40–320 ng/ml) in an iron-loaded or iron-free form. Changes in cell phenotype were analyzed by flow cytometry. NGAL stimulated expression of HLA-G on CD4+ T cells in a dose- and iron-dependent manner. Iron deficiency prevented NGAL-mediated effects, such that HLA-G expression was unaltered. Furthermore, NGAL treatment affected stimulation of regulatory T cells and in vitro expansion of CD4+ CD25+ FoxP3+ cells. An NGAL neutralizing antibody limited HLA-G expression and significantly decreased the percentage of CD4+ CD25+ FoxP3+ cells. Conclusions/Significance We provide in vitro evidence that NGAL is involved in cellular immunity. The potential role of NGAL as an immunomodulatory molecule is based on its ability to induce immune tolerance by upregulating HLA-G expression and expansion of T-regulatory cells in healthy donors. Future studies should further evaluate the role of NGAL in immunology and immunomodulation and its possible relationship to immunosuppressive therapy efficacy, tolerance induction in transplant patients, and other immunological disorders.

Outcome of pregnancy after organ transplantation: a retrospective survey in Italy

The number of women who decide to have a child after organ transplantation has increased. We determined the outcomes of 67 pregnancies of women who had undergone kidney, liver or heart transplantation. All recipients had been maintained on immunosuppressive therapy before and during pregnancy. Pregnancy complications at term were observed in 17 out of 67 women (25%), hypertension being the most frequent complication (16.17%). Two transplant rejections were reported. Sixty-eight infants were delivered (including one pair of twins); five women had two pregnancies at term. Twenty-eight miscarriages (29.2%) were recorded. Of these 68 babies (including the pair of twins), 40 (58.8%) were born at term and 28 (41.2%) before term. The babies were followed-up for 2 months to 13 years. According to our previous experience, our study shows that patients who have undergone organ transplantation can give birth to healthy infants as long as they are monitored accurately during pregnancy.

Incidence and predictors of postoperative atrial fibrillation in kidney transplant recipients.

Background Postoperative atrial fibrillation (POAF) is a complication of cardiothoracic and noncardiothoracic surgery. Kidney transplant recipients bear several known risk factors and may have a higher incidence of POAF. We retrospectively studied kidney and kidney/liver transplant recipients to estimate their POAF incidence and identify relevant risk factors. We also adapted a clinical score originally designed to predict thromboembolic risk in atrial fibrillation (AF; CHA2DS2-VASc) for assessing transplant patients. Methods We reviewed the clinical charts of kidney or kidney/liver transplant recipients from January 2005 to December 2008 at St. Orsola University Hospital Kidney Transplant Centre. Patients with and without POAF were compared on a number of clinical, laboratory, and instrumental data. Results The POAF incidence in kidney transplant recipients was 8.2%. Risk factors for POAF identified in univariate analyses included older recipient age, history of myocardial infarction, history of AF, liver/kidney transplantation, arterial stiffness, atherosclerotic plaques in the aorta or lower limbs, and diabetes mellitus. In a multivariate analysis, age, myocardial infarction history and combined liver/kidney transplantation were significant independent predictors of POAF. The modified CHA2DS2-VASc score proved to have a better predictive validity that the original CHA2DS2-VASc (area under the curve=0.71, 95% confidence interval=0.63–0.79 vs. area under the curve=0.62, 95% confidence interval=0.52–0.73, respectively). Conclusion AF is a notable complication of kidney, and particularly simultaneous liver/kidney, transplant surgery. Age, previous myocardial infarction, and simultaneous liver/kidney transplant independently predicted POAF. The modified CHA2DS2-VASc score could be useful to predict POAF risk in kidney transplant candidates.

Influence of the immunogenetic KIR and HLA systems on long-term renal transplant outcome

BACKGROUND Numerous studies have established the importance of innate immunity, particularly natural killer (NK) cells, in transplantation tolerance. NK cells express killer cell immunoglobulin-like receptors (KIRs) on their surface. By recognizing and binding major histocompatibility complex class I antigens, KIRs prevent autologous cell killing and promote lysis of non-self antigen-presenting cells. This study investigated the role of 16 KIR genes and donor-recipient KIR/HLA combinations on 5-year outcomes in a population of deceased donor kidney transplant recipients. MATERIAL/METHODS We genotyped 126 renal transplant patients and their donors for HLA A, B, C, DR, and KIR genes. Patients underwent standardized transplantation and immunosuppressive protocols and were followed-up for 5 years. Graft function was evaluated by serum creatinine level and glomerular filtration rate calculated using the 4-variable modification of diet in renal disease (MDRD) equation. RESULTS The presence of KIR2DS3 in the recipients was associated with better graft function indexes over time (p<0.05), but this effect was not confirmed by multivariate analysis. Conversely, the presence KIR2DS3 in the recipients combined with the presence of its HLA ligand in the donor had a detrimental effect on the trends of serum creatinine levels and eGFR trends, also confirmed by multivariate analysis. Kidney transplant recipients negative for the KIR2DL1 gene displayed higher creatinine levels after 5 years. Lastly, transplantation of HLA-A3/A11-negative donor kidneys into KIR3DL2-positive patients exerted a protective effect in terms of 5-years outcome (p<0.05). CONCLUSIONS The present study demonstrates an important role of the KIR immunogenetic system in the long-term immune response to kidney transplantation.

Reduction of Oxidative Damage Reflects a Better Kidney Transplantation Outcome

Background/Aims: DNA fragmentation is one of the typical features of apoptosis, frequently induced by oxidative stress. Increased oxidative stress is known to be related to several pathological processes. In this study, we assessed oxidative damage in the early follow-up period after kidney transplantation measuring DNA oxidation and fragmentation of mononuclear cells and the circulating levels of inflammatory cytokines. Methods: Blood samples from 30 kidney transplant recipients were collected before transplantation and after 2 days, 1 month and 6 months. Oxidative DNA fragmentation was measured by Comet Assay, whereas DNA oxidation was evaluated measuring 8-OHdG leukocyte levels. Serum IL-1β, IL-4, IL-6, IL-8, IL-10, IFN-γ and TNF-α were assayed using a multiplex ELISA analysis. Results: At 6 months after transplantation, a significant reduction in DNA fragmentation and IL-6 plasma levels was observed; DNA oxidation was higher in patients with a worse outcome, with delayed graft function and low nutritional status. We also found a correlation of IL-6 and IL-10 levels with DNA fragmentation and of IL-10 levels with DNA oxidation. Conclusion: Low levels of oxidation and apoptosis at 6 months after transplantation correlate with a better recovery of renal function in kidney allografts. The measurement of cytokine levels confirmed a reduction of inflammatory parameters within 6 months of follow-up.

Hormonal changes in hemodialysis

When chronic renal failure becomes advanced, the serum or tissue levels and/or the functions of most hormones are altered because of several interplaying mechanisms involving synthesis, transport, accumulation of inhibitors, abnormality of target organ responsiveness, and impaired renal clearance. As regular dialysis treatment is performed, most of these uremic hormone abnormalities are not reversed, and some of them may even get worse. The main endocrine derangements can be grouped as sexual hormone dysfunction, thyroid abnormalities, growth retardation, hormone-related disorders of metabolism, derangements of pressor substances, gastrointestinal peptide abnormalities, renal osteodystrophy, and anemia. The endocrine abnormalities play a major role as for the clinical and metabolic rehabilitation of dialysis patients.

Incidence of Late Deep Venous Thrombosis Among Renal Transplant Patients

BACKGROUND Kidney transplant recipients (KTRs) manifest hypercoagulable state that contributes to an increased incidence of deep vein thrombosis (DVT), not only early but also late in their course. KTRs display an imbalance of hemostatic mechanisms with a multifactorial rise in procoagulant factors, partly related to traditional risk factors and partly to transplantation. The aim of this study was to evaluate the incidence of first episodes of DVT among KTRs, focusing on risk factors. METHODS From 2008 to 2011, we evaluated 30 kidney transplant patients who ≥4 months there after transplantation developed DVT in the lower limbs only, lower limbs complicated by pulmonary embolism or retinal thrombosis. We analyzed causes of primary nephropathy, immunosuppressive regimen, post-transplantation infections, and erythrocytosis. DVT was diagnosed by color Doppler ultrasound or eye examination. RESULTS A significantly increased incidence of DVT was observed among patients receiving cyclosporine or cyclosporine + mammalian target of rapamycin inhibitors, affected by polycystic kidney diseases, systemic lupus erythematosus or nephrotic syndrome, or displaying rapid and/or excessive correction of hematocrit values. DVT was not significantly related to an acute infection (cytomegalovirus) or to the prior dialysis modality. CONCLUSIONS Hypercoagulability is a multifactorial condition in KTRs, representing a severe complication in stable patients. Prevention may consist of either accurate pretransplantation screening for thrombophilia or identification of patients at higher DVT risk.

Multicenter Prospective Study for Laboratory Diagnosis of HHV8 Infection in Solid Organ Donors and Transplant Recipients and Evaluation of the Clinical Impact After Transplantation

Background We performed serological and molecular pretransplant screening in solid organ transplant (SOT) donors and recipients in north central Italy and a surveillance program for human herpesvirus 8 (HHV8) infection after transplant, aiming to establish an optimal management of HHV8 infection in SOT recipients. Methods For pretransplant HHV8 screening in both donors and recipients, 6 serological (4 indirect immunofluorescent assays [IFA] and 2 enzyme-linked immunosorbent assays—both HHV8 lytic and latent antigen based) and 2 molecular assays were used. A reference standard to identify HHV8-positive patients was defined by at least 2 positive assays. All transplant patients at risk to develop HHV8-related disease underwent virological posttransplant monitoring by quantitative real-time polymerase chain reaction (PCR) assay. Results Human herpesvirus 8 seroprevalence was 4% (10/249) in donors and 18% (93/517) in organ recipients. The best performance was obtained by 2 lytic antigen-based IFAs that showed almost perfect agreement to the reference standard (0.943 and 0.931 Cohen kappa). Human herpesvirus 8–DNA was detected in 6.8% and 2.9% of HHV8-seropositive donor samples by in-house nested PCR and quantitative real-time PCR assays, respectively. After transplant, 3 (25%) of 12 HHV8-mismatch patients (seropositive donor/seronegative recipient) developed a primary infection, one of whom developed a lethal nonmalignant illness. Two of 93 HHV8-seropositive recipients (2.1%) had viral replication in posttransplant period, one of whom developed Kaposi sarcoma. Conclusions Serological assays, specifically lytic IFAs, were the best methodological approach to identify HHV8-infected SOT donors and recipients. A very low incidence (1.9%) of posttransplant HHV8-related disease was observed.