Isabella Guzzo

Inhibition of B-Cell Proliferation and Antibody Production by Mesenchymal Stromal Cells Is Mediated by T Cells

Bone marrow (BM)-derived mesenchymal stromal cells (MSCs), endowed with immunosuppressive and anti-inflammatory properties, represent a promising tool in immunoregulatory and regenerative cell therapy. Clarifying the interactions between MSCs and B-lymphocytes may be crucial for designing innovative MSC-based strategies in conditions in which B cells play a role, including systemic lupus erythematosus (SLE) and rejection of kidney transplantation. In this study, we show that, both in healthy subjects and in patients, in vitro B-cell proliferation, plasma-cell differentiation, and antibody production are inhibited by BM-derived MSCs when peripheral blood lymphocytes are stimulated with CpG, but not when sorted B cells are cultured with MSCs+CpG. Inhibition is restored in CpG+MSC cocultures when sorted T cells are added to sorted B cells, suggesting that this effect is mediated by T cells, with both CD4(+) and CD8(+) cells playing a role. Moreover, cell-cell contact between MSCs and T cells, but not between MSCs and B cells, is necessary to inhibit B-cell proliferation. Thus, the presence of functional T cells, as well as cell-cell contact between MSCs and T cells, are crucial for B-cell inhibition. This information can be relevant for implementing MSC-based therapeutic immune modulation in patients in whom T-cell function is impaired.

Use of rituximab in focal glomerulosclerosis relapses after renal transplantation.

Background. Focal and segmental glomerulosclerosis (FSGS) accounts for more than 10% of all cases of renal diseases leading to renal failure in children. After renal transplantation, 20% to 40% of FSGS relapse, frequently leading to renal loss. Plasmapheresis is considered the first option to treat relapses by several authors but is often ineffective. The anti-CD20 monoclonal antibody rituximab has been proposed as a possible treatment. Methods. We reviewed the effect of rituximab in seven children or young adults with pretransplant FSGS and relapse of proteinuria after transplantation who did not respond to intensive plasmapheresis. Results. After treatment, urine protein disappeared in three patients, was reduced by 70% in one patient and by 50% in one patient. No response was observed in one patient who had a quick deterioration of renal function and reached end-stage renal failure after 3 months. One additional patient developed a severe reaction a few minutes after the start of the first rituximab infusion. Conclusion. Rituximab is a possible option to treat some resistant cases of FSGS with relapsing proteinuria after transplantation. It is important that therapy is started early after evidence of failure of plasmapheresis, before sclerosis develops in the glomeruli. The response to treatment can occur after several months. During the follow-up period, CD19 cells should be monitored carefully, and additional rituximab infusions considered to maintain B-cell depletion.

Three-yr safety and efficacy of everolimus and low-dose cyclosporine in de novo pediatric kidney transplant patients

The three yr results of a multicenter trial in de novo pediatric KT treated with a proliferative signal inhibitor and low dose CNI are presented. Thirty‐seven children (9.1 ± 5 yr old) received basiliximab, cyclosporine A (CyA C2:1400 ng/mL), (MMF C0:1.5–3 μg/mL), and prednisone. Three wk later everolimus was started (C0:5–10 ng/mL), CyA was reduced (C2:600 ng/mL after 90 days 300 ng/mL), and MMF discontinued. During the three‐yr period patient and graft survivals were 96%. One patient died for causes unrelated to the immunosuppression. Cumulative acute rejection rate including protocol and indication biopsies was 21.9%. None of the patients had signs of chronic humoral rejection. Incidence of dnDSA was 5%, 11%, and 22% at one, two, and three yr post‐transplant, respectively. Mean glomerular filtration rate measured at one yr and three yr post‐transplant was 105.5 ± 31 and 110.7 ± 27 mL/min/1.73 m2, respectively. A growth velocity of 7.7 ± 6.7 cm/yr was achieved with positive catch‐up growth. No malignancy or post‐transplant lymphoproliferative diseases were diagnosed. In conclusion, the treatment based on basiliximab induction, everolimus, low‐dose cyclosporine, and low‐dose prednisone leads to good long‐term efficacy in de novo pediatric KT recipients.

Post-transplant recurrence of atypical hemolytic uremic syndrome in a patient with thrombomodulin mutation

HUS is characterized by hemolytic anemia, thrombocytopenia, and acute renal failure. While “typical” HUS is usually associated with Shiga toxin‐producing Escherichia coli infections and recovers in the majority of cases, aHUS is caused by mutations of complement components or antibodies against CFH leading to uncontrolled activation of alternative complement pathway and often to ESRD. Recently, THBD gene mutations have been reported in aHUS. Theoretically, the risk of disease recurrence after renal transplantation should be low because THBD is primarily a membrane‐bound protein expressed by endothelial cells; however, a small proportion of THBD is present as a soluble form in plasma. We report the case of a 19‐yr‐old man with aHUS secondary to a THBD mutation that relapsed twice after two renal transplantations performed 12 yr apart. Despite successful control of HUS with plasma exchange and eculizumab after the second transplantation, the graft was ultimately lost due to severe steroid‐resistant cellular rejection. The present report suggests that THBD mutations may favor‐relapse of aHUS after renal transplantation.

Cytomegalovirus Infection in Pediatric Renal Transplantation and the Impact of Chemoprophylaxis With (Val-)Ganciclovir

Background Cytomegalovirus (CMV) replication and disease, with its associated morbidity and poor transplant outcome, represents a serious threat to transplant recipients. The pediatric kidney transplant population is at a particularly increased risk of CMV infection. Methods We therefore analyzed CMV epidemiology in a large cohort of pediatric renal transplant recipients (n = 242) and assessed the impact of antiviral chemoprophylaxis with valganciclovir (VGCV) or ganciclovir (GCV) on CMV replication and morbidity. Results While antiviral chemoprophylaxis with VGCV or GCV in patients with a high (D+/R−) or intermediate (D+/R+) CMV risk (n = 82) compared to preemptive therapy (n = 47) had no significant effect on the incidence of CMV syndrome or tissue-invasive disease, chemoprophylaxis was associated with a better preservation of transplant function at 3 years posttransplant (loss of estimated glomerular filtration rate in the chemoprophylaxis cohort, 16.0 ± 3.4 vs. 30.1 ± 4.7 mL/min per 1.73 m2 in the preemptive therapy cohort, P < 0.05).CMV replication was associated with a more pronounced decline of graft function (difference in estimated glomerular filtration rate of 9.6 mL/min per 1.73 m2 at 3 years) compared to patients without CMV replication. However, patients undergoing VGCV or GCV chemoprophylaxis had more leukocytopenia. Conclusion Antiviral chemoprophylaxis with VGCV or GCV in recipients with a high or moderate CMV risk is associated with a better preservation of transplant function. Hence, the prevention of CMV replication in this patient population has the potential to improve transplant outcome.

Premature ageing of the immune system relates to increased anti‐lymphocyte antibodies (ALA) after an immunization in HIV‐1‐infected and kidney‐transplanted patients

Low‐affinity immunoglobulin (Ig)G with potential autoreactivity to lymphocytes and hypergammaglobulinaemia have been described previously in HIV‐1‐infected patients. Whether such antibodies increase after challenging the immune system, for example with an immunization, is not known. In the present study, the modulation of antibodies with low affinity and potential autoreactivity was evaluated after 2012–13 seasonal flu vaccination with a simple empirical laboratory test measuring the titres of anti‐lymphocyte antibodies (ALA) in two different models of secondary immunodeficiency: HIV‐1 vertically infected patients (HIV) and patients treated with immunosuppressive therapies after kidney transplantation (KT) compared to healthy individuals (HC). In parallel, the activation status of B cells and their degree of immune senescence was evaluated by measuring the B cell interleukin (IL)‐21R expression/plasma IL‐21 levels and the frequencies of mature‐activated (MA) and double‐negative (DN) B cells. A significant increase of ALA titres was observed after vaccination in HIV and KT but not in HC, and this correlated directly with the frequencies of both MA and DN and inversely with the B cell IL‐21R expression. This suggests that the quality of an immune response triggered by flu vaccination in HIV and KT may depend upon the activation status of B cells and on their degree of immune senescence. Further investigations are needed to verify whether high frequencies of MA and DN may also relate to increase autoimmunity after immunization in high‐risk populations.

Impact of Everolimus and Low-Dose Cyclosporin on Cytomegalovirus Replication and Disease in Pediatric Renal Transplantation

In order to investigate the hypothesis that the mammalian target of rapamycin inhibitor everolimus (EVR) shows anticytomegalovirus (CMV) activity in pediatric patients, we analyzed the impact of EVR‐based immunosuppressive therapy on CMV replication and disease in a large cohort (n = 301) of pediatric kidney allograft recipients. The EVR cohort (n = 59), who also received low‐dose cyclosporin, was compared with a control cohort (n = 242), who was administered standard‐dose cyclosporin or tacrolimus and an antimetabolite, mostly mycophenolate mofetil (91.7%). Multivariate analysis revealed an 83% lower risk of CMV replication in the EVR cohort than in the control cohort (p = 0.005). In CMV high‐risk (donor+/recipient−) patients (n = 88), the EVR‐based regimen was associated with a significantly lower rate of CMV disease (0% vs. 14.3%, p = 0.046) than the standard regimen. In patients who had received chemoprophylaxis with (val‐)ganciclovir (n = 63), the CMV‐free survival rates at 1 year and 3 years posttransplant (100%) were significantly (p = 0.015) higher in the EVR cohort (n = 15) than in the control cohort (n = 48; 1 year, 75.0%; 3 years, 63.3%). Our data suggest that in pediatric patients at high risk of CMV, an EVR‐based immunosuppressive regimen is associated with a lower risk of CMV disease than a standard‐dose calcineurin inhibitor–based regimen.

B-sides serologic markers of immunogenicity in kidney transplanted patients: report from 2012-2013 flu vaccination experience.

Background Safety and immunogenicity data of seasonal influenza vaccination in transplanted patients (Tps) are controversial. Preexisting cross-reactive antibodies generated by repeated vaccination with drift variant strains could bias interpretation of immunogenicity data in Tp. Methods The unadjuvanted 2012–2013 seasonal influenza vaccine was administered to 81 kidney Tps being routinely vaccinated against influenza and 23 healthy controls (HCs). Immunogenicity was evaluated by both strain-specific antibody responses with standard hemagglutination inhibition assay and by memory B-cell enzyme-linked immunosorbent spot. Safety was also evaluated by measuring anti–human leukocyte antigen (HLA) antibodies. Results The majority of Tps were seroprotected before vaccination (81.5%, 81.5%, and 43.2% vs. 47.8%, 34.8%, and 30.4% in HC for H1N1, H3N2, and B strain, respectively) resulting into lower seroconversion rates (P⩽0.01) as compared with HC (40.7%, 39.5%, and 54.3% vs. 73.9%, 82.6%, and 65.2% for H1N1, H3N2, and B strain, respectively). An inverse correlation was found between seroconversion rates and number of previous vaccinations in Tps. On the contrary, similar increase in the frequencies of strain-specific memory B cells were detected by B-cell enzyme-linked immunosorbent spot in both Tps and HCs after vaccination. No serious adverse events have been reported. Donor-specific HLA antibodies increased in two patients after vaccination, and de novo anti-HLA antibodies were identified in two additional patients (non–donor-specific HLA antibodies). Conclusion This report on safety and immunogenicity of the seasonal unadjuvanted 2012–2013 flu vaccination suggests that evaluating immunogenicity of influenza vaccination exclusively by hemagglutination inhibition assay may be misleading in individuals receiving yearly seasonal vaccines. Further investigations are required to understand the relation between vaccination and anti-HLA antibody development.

Circuit lifespan during continuous renal replacement therapy: children and adults are not equal.

In the field of continuous renal replacement therapy (CRRT), session length, downtime and dose require detailed research, which will provide information important in relation to prescription, anticoagulation and circuit material choice (membrane type and size, vascular access site and size). In particular, it appears that many of the data currently existing in the literature and accepted regarding CRRT prescription and delivery in critically ill adult patients are not strictly applicable to the paediatric setting. Furthermore, many of the available paediatric studies are small, retrospective or underpowered. In paediatric CRRT, epidemiological investigations and prospective trials to investigate practical aspects of extracorporeal therapies are welcome and urgently needed.

Acute kidney transplant rejection mediated by angiotensin II type 1 receptor antibodies in a pediatric hyperimmune patient

BackgroundSeveral cases of severe antibody-mediated rejection (AMR) secondary to antibodies against the angiotensin II type 1 receptor (AT1R-Ab) have been described with variable outcome.Case-Diagnosis/TreatmentWe report the case of a 13-year-old boy whose first kidney transplant failed due to steroid-resistant acute cellular rejection, with the subsequent development of sensitization. He received a second kidney transplant which was complicated by early humoral rejection, with weakly positive staining for the complement degradation product C4d. Test results were negative for donor-specific antibodies against human leukocyte antigens (HLA-DSA) and MHC class I-related chain A (MICA) but positive for AT1R-Ab. Retrospective testing of the sera collected during the first kidney transplant was also positive for AT1R-Ab. We therefore hypothesized that the failure of the first transplant was secondary to the same cause. Losartan was immediately introduced into the therapeutic regimen, and the patient showed an excellent clinical and histological recovery.ConclusionsTesting for AT1R-Ab in any hypertensive patient with acute rejection who tests negative or weakly positive for C4d and negative for HLA-DSA and who is refractory to therapy is highly advisable. Pre-transplant AT1R-Ab may be indicative of the outcome in patients whose first transplant failed. Prompt initiation of treatment with losartan—immediately after transplantation in patients with pre-existing AT1R-Ab—should be encouraged.