Attenuation of Torque teno viral load over time in kidney transplantation recipients treated with calcineurin inhibitors is mitigated after conversion to belatacept.

Torque Teno Virus (TTV) is a non-pathogenic anellovirus, highly prevalent in healthy populations. Variations in its viral load have been associated with states of diminished immunity, as occurs after organ transplantation. It is hypothesized that TTV-load might be used as a diagnostic tool to guide prescription and dosing of immunosuppressive drugs. Not much is known about the effects of combined immunosuppressive drugs on TTV replication in renal transplantation. Belatacept was introduced to counter side-effects of calcineurin inhibitors (CNI). It was never widely adopted, mainly because its association with increased risk of rejection. To investigate the differential effects of a regimen based on calcineurin inhibitors versus belatacept on TTV-loads, we measured TTV-levels in 105 patients from two randomized controlled trials in kidney transplant recipients (KTRs). We observed that time after transplantation was inversely related to TTV-levels of patients that remained on a CNI-containing regime, whereas this decline over time was diminished after conversion to belatacept. In addition, a correlation with tacrolimus-trough levels and age were found. Our study is the first report on the impact of conversion from CNI to belatacept on TTV-levels in KTR. In conclusion, the time-related decline in TTV-levels is mitigated after conversion from CNI to belatacept.

Conversion from calcineurin inhibitors to belatacept-based immunosuppressive therapy skews terminal proliferation of non-classical monocytes and lowers lymphocyte counts.

Belatacept, a modified form of CTLA-Ig that blocks CD28-mediated co-stimulation of T cells, is an immune-suppressant that can be used as an alternative to calcineurin inhibitors (CNIs). In kidney transplant recipients, belatacept has been associated with improved renal function and reduced cardiovascular toxicity. Monocytes as well as T-lymphocytes play causal roles in the pathophysiology of atherosclerotic disease. We hypothesized that the beneficial impact of the use of belatacept over CNIs on cardiovascular risk could be partly explained by the impact of belatacept therapy on these circulating leukocytes. Hence, we phenotyped circulating leukocytes in transplanted patients with a stable renal function that were randomized between either continuation of CNI or conversion to belatacept in two international studies in which we participated. In 41 patients, we found that belatacept-treated patients consistently showed lower numbers of B-lymphocytes, T-lymphocytes as well as CD14-negative monocytes (CD14NM), especially in non-diabetic patients. Our observation that this decrease was associated to plasma concentrations of TNFα is consistent with a model where CD14NM-production of TNFα is diminished by belatacept-treatment, due to effects on the antigen-presenting cell compartment.

Torque teno virus load as marker of rejection and infection in solid organ transplantation - A systematic review and meta-analysis.

Balancing immunosuppression to prevent rejection in solid organ transplant (SOT) recipients remains challenging. Torque teno virus (TTV), a commensal non-pathogenic virus, has been proposed as marker of functional immunity: higher loads correspond to over-immunosuppression, and lower loads to under-immunosuppression. This review offers an overview of the current evidence of the association between TTV-load and infection and rejection after SOT. A systematic literature search strategy, deposited in the PROSPERO registry, resulted in 548 records. After screening, 23 original and peer-reviewed articles were assessed investigating the association between TTV-load, infection and/or rejection in SOT. The Quality in Prognostic Studies (QUIPS)-tool was used to assess the risk of bias. Meta-analysis with random-effects was performed on results with similar outcomes and exposure measures. Most of the included studies involved retrospective cohorts in which the TTV-load was measured longitudinally, within the first 2 years post-transplantation. Infection outcomes differed between studies and included viral, bacterial, parasitic and fungal infections. Rejection was defined by biopsy confirmation or initiation of rejection treatment. Twelve out of 16 studies reported an association between high TTV-load and infections, whereas 13 out of 15 reported an association between low TTV-load and rejection. Meta-analysis showed an increased risk of infection (OR: 1.16, 95% CI: 1.03-1.32; HR: 1.05, 95% CI: 0.97-1.14) and a decreased risk of rejection (OR: 0.90, 95% CI: 0.87-0.94; HR: 0.74, 95% CI: 0.71-0.76) per 1 log TTV-load increase. The qualitative assessment showed varying risks of bias in the included studies. This systematic review and meta-analysis indicates that blood TTV-load measured within the first 2 years after SOT is associated with the risk of infection or allograft rejection, although substantial risk of bias in the studies included warrant cautious interpretation. The results in this review provide a rationale for larger, prospective, studies into TTV as marker of infection and rejection after SOT.

Presence of CD163+ macrophages in DCD kidneys with high DGF reduces the risk for acute cellular rejection in 6 months after kidney transplantation.

Acute cellular rejection (ACR) occurs in 10% of renal allograft recipients and is characterized by leukocyte infiltration as observed in needle biopsies. ACR onset is subject to several risk factors, including delayed graft function (DGF). As the impact of DGF on the etiology of ACR remains unclear, this study analyzed the association between presence of leukocyte subsets and ACR onset, in DCD kidney biopsies with extensive DGF following transplantation. Immunohistochemical analysis of protocol biopsies taken 10 days after kidney transplantation revealed that patients with high levels of renal CD163+ macrophages have a decreased risk (OR = 0.021, P = 0.008) for ACR in the first 6 months after transplantation. In pre-transplant biopsies of a comparable DCD cohort, with >80% DGF, presence of donor CD163+ macrophages showed no effect on ACR risk. Therefore, leukocyte infiltrate present during the inflammatory response at the time of DGF may contain anti-inflammatory macrophages that exert a protective effect against ACR development.

A novel method for engineering autologous non-thrombogenic in situ tissue-engineered blood vessels for arteriovenous grafting.

The durability of prosthetic arteriovenous (AV) grafts for hemodialysis access is low, predominantly due to stenotic lesions in the venous outflow tract and infectious complications. Tissue engineered blood vessels (TEBVs) might offer a tailor-made autologous alternative for prosthetic grafts. We have designed a method in which TEBVs are grown in vivo, by utilizing the foreign body response to subcutaneously implanted polymeric rods in goats, resulting in the formation of an autologous fibrocellular tissue capsule (TC). One month after implantation, the polymeric rod is extracted, whereupon TCs (length 6 cm, diameter 6.8 mm) were grafted as arteriovenous conduit between the carotid artery and jugular vein of the same goats. At time of grafting, the TCs were shown to have sufficient mechanical strength in terms of bursting pressure (2382 ±â€¯129 mmHg), and suture retention strength (SRS: 1.97 ±â€¯0.49 N). The AV grafts were harvested at 1 or 2 months after grafting. In an ex vivo whole blood perfusion system, the lumen of the vascular grafts was shown to be less thrombogenic compared to the initial TCs and ePTFE grafts. At 8 weeks after grafting, the entire graft was covered with an endothelial layer and abundant elastin expression was present throughout the graft. Patency at 1 and 2 months was comparable with ePTFE AV-grafts. In conclusion, we demonstrate the remodeling capacity of cellularized in vivo engineered TEBVs, and their potential as autologous alternative for prosthetic vascular grafts.

Source and Relevance of the BK Polyomavirus Genotype for Infection After Kidney Transplantation.

BACKGROUND: BK polyomavirus (BKPyV)-associated nephropathy (BKPyVAN) is a major threat for kidney transplant recipients (KTRs). The role of specific BKPyV genotypes/serotypes in development of BKPyVAN is poorly understood. Pretransplantation serotyping of kidney donors and recipients and posttransplantation genotyping of viremic recipients, could reveal the clinical relevance of specific BKPyV variants. METHODS: A retrospective cohort of 386 living kidney donor-recipient pairs was serotyped before transplantation against BKPyV genotype I-IV viral capsid protein 1 antigen, using a novel BKPyV serotyping assay. Replicating BKPyV isolates in viremic KTRs after transplantation were genotyped using real-time polymerase chain reaction and confirmed by means of sequencing. BKPyV serotype and genotype data were used to determine the source of infection and analyze the risk of viremia and BKPyVAN. RESULTS: Donor and recipient BKPyV genotype and serotype distribution was dominated by genotype I (>80%), especially Ib, over II, III and IV. Donor serotype was significantly correlated with the replicating genotype in viremic KTRs (P < .001). Individual donor and recipient serotype, serotype (mis)matching and the recipient replicating BKPyV genotype were not associated with development of viremia or BKPyVAN after transplantation. CONCLUSIONS: BKPyV donor and recipient serotyping and genotyping indicates the donor origin of replicating BKPyV in viremic KTRs but provides no evidence for BKPyV genotype-specific virulence.

Parathyroidectomy versus cinacalcet for tertiary hyperparathyroidism; a retrospective analysis.

INTRODUCTION: Tertiary hyperparathyroidism (tHPT), i.e., persistent HPT after kidney transplantation, affects 17-50% of transplant recipients. Treatment of tHPT is mandatory since persistently elevated PTH concentrations after KTx increase the risk of renal allograft dysfunction and osteoporosis. The introduction of cinacalcet in 2004 seemed to offer a medical treatment alternative to parathyroidectomy (PTx). However, the optimal management of tHPT remains unclear. METHODS: A retrospective analysis was performed on patients receiving a kidney transplantation (KT) in two academic centers in the Netherlands. Thirty patients undergoing PTx within 3 years of transplantation and 64 patients treated with cinacalcet 1 year after transplantation for tHPT were included. Primary outcomes were serum calcium and PTH concentrations 1 year after KT and after PTx. RESULTS: Serum calcium normalized in both the cinacalcet and the PTx patients. PTH concentrations remained above the upper limit of normal (median 22.0 pmol/L) 1 year after KT, but returned to within the normal range in the PTx group (median 3.7 pmol/L). Side effects of cinacalcet were difficult to assess; minor complications occurred in three patients. Re-exploration due to persistent tHPT was performed in three (10%) patients. CONCLUSION: In patients with tHPT, cinacalcet normalizes serum calcium, but does not lead to a normalization of serum PTH concentrations. In contrast, PTx leads to a normalization of both serum calcium and PTH concentrations. These findings suggest that PTx is the treatment of choice for tHPT.

Abdominal aortic calcification on a plain X-ray and the relation with significant coronary artery disease in asymptomatic chronic dialysis patients.

BACKGROUND: Coronary artery disease (CAD) is common in asymptomatic chronic dialysis patients and plays an important role in their poor survival. Early identification of these high-risk patients could improve treatment and reduce mortality. Abdominal aortic calcification (AAC) has previously been associated with CAD in autopsy studies. Since the AAC can be quantified easily using a lateral lumbar X-ray we hypothesized that the extent of AAC as assessed on a lateral lumbar X-ray might be predictive of the presence of significant CAD in dialysis patients. METHODS: All patients currently enrolled in the ICD2 trial without a history of CABG or a PCI with stent implantation were included in this study. All patients underwent CT-angiography (CTA) and a lateral X-ray of the abdomen. AAC on X-ray was quantified using a previously validated scoring system whereupon the association between AAC and the presence of significant CAD was assessed. RESULTS: A total of 90 patients were included in this study (71% male, 67 ± 7 years old). Forty-six patients were found to have significant CAD. AAC-score was significantly higher in patients with CAD (10.1 ± 4.9 vs 6.3 ± 4.6 (p < 0.05). Multivariate regression analysis revealed that AAC score is an independent predictor for the presence of CAD with a 1,2 fold higher risk per point increase (p < 0.01). The AAC score has a sensitivity of 85% and a specificity of 57% for the presence of significant CAD. CONCLUSION: This study shows that abdominal aortic calcification as assessed on a lateral lumbar X-ray is predictive for the presence of significant coronary artery disease in asymptomatic dialysis patients. This simple, non-invasive and cheap screening method could contribute to early identification of patients eligible for further screening of CAD. TRIAL REGISTRATION: NTR948 , registered 10-4-2007 ; ISRCTN20479861 , registered 2-5-2007.

Pretransplantation Donor-Recipient Pair Seroreactivity Against BK Polyomavirus Predicts Viremia and Nephropathy After Kidney Transplantation.

Kidney transplant donors are not currently implicated in predicting BK polyomavirus (BKPyV) infection in kidney transplant recipients. It has been postulated, however, that BKPyV infection originates from the kidney allograft. Because BKPyV seroreactivity correlates with BKPyV replication and thus might mirror the infectious load, we investigated whether BKPyV seroreactivity of the donor predicts viremia and BKPyV-associated nephropathy (BKPyVAN) in the recipient. In a retrospective cohort of 407 living kidney donor-recipient pairs, pretransplantation donor and recipient sera were tested for BKPyV IgG levels and correlated with the occurrence of recipient BKPyV viremia and BKPyVAN within 1 year after transplantation. Donor BKPyV IgG level was strongly associated with BKPyV viremia and BKPyVAN (p < 0.001), whereas recipient BKPyV seroreactivity showed a nonsignificant inverse trend. Pairing of high-BKPyV-seroreactive donors with low-seroreactive recipients resulted in a 10-fold increased risk of BKPyV viremia (hazard ratio 10.1, 95% CI 3.5-29.0, p < 0.001). In multivariate analysis, donor BKPyV seroreactivity was the strongest pretransplantation factor associated with viremia (p < 0.001) and BKPyVAN (p = 0.007). The proportional relationship between donor BKPyV seroreactivity and recipient infection suggests that donor BKPyV seroreactivity reflects the infectious load of the kidney allograft and calls for the use of pretransplantation BKPyV serological testing of (potential) donors and recipients.

Safety and long-term effects of renal denervation: Rationale and design of the Dutch registry.

BACKGROUND: Percutaneous renal denervation (RDN) has recently been introduced as a treatment for therapy-resistant hypertension. Also, it has been suggested that RDN may be beneficial for other conditions characterised by increased sympathetic nerve activity. There are still many uncertainties with regard to efficacy, safety, predictors for success and long-term effects. To answer these important questions, we initiated a Dutch RDN registry aiming to collect data from all RDN procedures performed in the Netherlands. METHODS: The Dutch RDN registry is an ongoing investigator-initiated, prospective, multicentre cohort study. Twenty-six Dutch hospitals agreed to participate in this registry. All patients who undergo RDN, regardless of the clinical indication or device that is used, will be included. Data are currently being collected on eligibility and screening, treatment and follow-up. RESULTS: Procedures have been performed since August 2010. At present, data from 306 patients have been entered into the database. The main indication for RDN was hypertension (n = 302, 99%). Patients had a mean office blood pressure of 177/100 (±29/16) mmHg with a median use of three (range 0-8) blood pressure lowering drugs. Mean 24-hour blood pressure before RDN was 157/93 (±18/13) mmHg. RDN was performed with different devices, with the Simplicity™ catheter currently used most frequently. CONCLUSION: Here we report on the rationale and design of the Dutch RDN registry. Enrolment in this investigator-initiated study is ongoing. We present baseline characteristics of the first 306 participants.

Elastin is a key regulator of outward remodeling in arteriovenous fistulas.

OBJECTIVES: Maturation failure is the major limitation of arteriovenous fistulas (AVFs) as hemodialysis access conduits. Indeed, 30-50% of AVFs fail to mature due to intimal hyperplasia and insufficient outward remodeling. Elastin has emerged as an important determinant of vascular remodeling. Here the role of elastin in AVF remodeling in elastin haplodeficient (eln(+/-)) mice undergoing AVF surgery has been studied. METHODS: Unilateral AVFs between the branch of the jugular vein and carotid artery in an end to side manner were created in wild-type (WT) C57BL/6 (n = 11) and in eln(+/-) mice (n = 9). Animals were killed at day 21 and the AVFs were analyzed histologically and at an mRNA level using real-time quantitative polymerase chain reaction. RESULTS: Before AVF surgery, a marked reduction in elastin density in the internal elastic lamina (IEL) of eln(+/-) mice was observed. AVF surgery resulted in fragmentation of the venous internal elastic lamina in both groups while the expression of the tropoelastin mRNA was 53% lower in the eln(+/-) mice than in WT mice (p < .001). At 21 days after AVF surgery, the circumference of the venous outflow tract of the AVF was 21% larger in the eln(+/-) mice than in the WT mice (p = .037), indicating enhanced outward remodeling in the eln(+/-) mice. No significant difference in intimal hyperplasia was observed. The venous lumen of the AVF in the eln(+/-) mice was 53% larger than in the WT mice, although this difference was not statistically significant (eln(+/-), 350,116 ± 45,073 µm(2); WT, 229,405 ± 40,453 µm(2); p = .064). CONCLUSIONS: In a murine model, elastin has an important role in vascular remodeling following AVF creation, in which a lower amount of elastin results in enhanced outward remodeling. Interventions targeting elastin degradation might be a viable option in order to improve AVF maturation.

Circulating microRNAs associate with diabetic nephropathy and systemic microvascular damage and normalize after simultaneous pancreas-kidney transplantation.

Because microvascular disease is one of the most important drivers of diabetic complications, early monitoring of microvascular integrity may be of clinical value. By assessing profiles of circulating microRNAs (miRNAs), known regulators of microvascular pathophysiology, in healthy controls and diabetic nephropathy (DN) patients before and after simultaneous pancreas-kidney transplantation (SPK), we aimed to identify differentially expressed miRNAs that associate with microvascular impairment. Following a pilot study, we selected 13 candidate miRNAs and determined their circulating levels in DN (n = 21), SPK-patients (n = 37), healthy controls (n = 19), type 1 diabetes mellitus patients (n = 15) and DN patients with a kidney transplant (n = 15). For validation of selected miRNAs, 14 DN patients were studied longitudinally up to 12 months after SPK. We demonstrated a direct association of miR-25, -27a, -126, -130b, -132, -152, -181a, -223, -320, -326, -340, -574-3p and -660 with DN. Of those, miR-25, -27a, -130b, -132, -152, -320, -326, -340, -574-3p and -660 normalized after SPK. Importantly, circulating levels of some of these miRNAs tightly associate with microvascular impairment as they relate to aberrant capillary tortuosity, angiopoietin-2/angiopoietin-1 ratios, circulating levels of soluble-thrombomodulin and insulin-like growth factor. Taken together, circulating miRNA profiles associate with DN and systemic microvascular damage, and might serve to identify individuals at risk of experiencing microvascular complications, as well as give insight into underlying pathologies.

The dialysis procedure as a trigger for atrial fibrillation: new insights in the development of atrial fibrillation in dialysis patients.

AIMS: Atrial fibrillation (AF) is common in dialysis patients and is associated with increased morbidity and mortality. The pathophysiology may be related to common risk factors for both AF and renal disease or to dialysis-specific factors. The purpose of this study was to determine whether and how AF onset relates to the dialysis procedure itself. METHODS: All dialysis patients enrolled in the implantable cardioverter defibrillator-2 (ICD-2) trial until January 2012, who were implanted with an ICD, were included in this study. Using the ICD remote monitoring function, the exact time of onset of all AF episodes was registered. Subsequently, this was linked to the timing of dialysis procedures. RESULTS: For the current study, a total of 40 patients were included, follow-up was 28 ± 16 months, 80% male, 70 ± 8 years old. A total of 428 episodes of AF were monitored in 14 patients. AF onset was more frequent on the days of haemodialysis (HD) (p<0.001) and specifically increased during the dialysis procedure itself (p=0.04). Patients with AF had a larger left atrium (p<0.001) and a higher systolic blood pressure before and after HD (p<0.001). CONCLUSION: This study provides insight in the exact timing of AF onset in relation to the dialysis procedure itself. In HD patients, AF occurred significantly more often on a dialysis day and especially during HD. These findings might help to elucidate some aspects of the pathophysiology of AF in dialysis patients and could facilitate early detection of AF in these high-risk patients.

Activated platelets correlate with mobilization of naïve CD34(+) cells and generation of CD34(+) /KDR(+) cells in the circulation. A meta-regression analysis.

BACKGROUND: Bone marrow-derived circulating CD34(+) progenitor cells participate in remodeling and repair of the vasculature. Coexpression of the kinase-insert domain-containing receptor (KDR) has been proposed to identify the regenerative capacity. Recently, we provided evidence that the major fraction of circulating CD34(+) /KDR(+) cells is not mobilized from bone marrow, but is generated at sites of vascular injury through interaction with platelets. OBJECTIVES: To determine the relationship between platelet activation, the recruitment of naïve CD34(+) cells and the generation of CD34(+) /KDR(+) progenitor cells in a broad range of (patho)physiologic conditions, a detailed meta-regression analysis was conducted. METHODS/RESULTS: Twenty-eight conditions were found in which the numbers of CD34(+) and/or CD34(+) /KDR(+) cells and the levels of soluble P-selectin, as a marker for in vivo platelet activation, were documented. To combine heterogeneous data from 214 selected articles, results were standardized to a uniform scale by calculating standardized mean differences (SMDs) obtained from patient and control cohorts. Subsequently, a random-effects meta-regression analysis was performed on pooled SMDs. CONCLUSIONS: Our systemic survey supports a model in which activated platelets are a determinant for mobilization of CD34(+) cells from the bone marrow and the generation of CD34(+) /KDR(+) cells in the circulation.

Microvascular damage in type 1 diabetic patients is reversed in the first year after simultaneous pancreas-kidney transplantation.

Simultaneous pancreas-kidney transplantation (SPK) is an advanced treatment option for type 1 diabetes mellitus (DM) patients with microvascular disease including nephropathy. Sidestreamdarkfield (SDF) imaging has emerged as a noninvasive tool to visualize the human microcirculation. This study assessed the effect of SPK in diabetic nephropathy (DN) patients on microvascular alterations using SDF and correlated this with markers for endothelial dysfunction. Microvascular morphology was visualized using SDF of the oral mucosa in DN (n = 26) and SPK patients (n = 38), healthy controls (n = 20), DM1 patients (n = 15, DM ≥ 40 mL/min) and DN patients with a kidney transplant (KTx, n = 15). Furthermore, 21 DN patients were studied longitudinally up to 12 months after SPK. Circulating levels of angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2) and soluble thrombomodulin (sTM) were measured using ELISA. Capillary tortuosity in the DN (1.83 ± 0.42) and DM ≥ 40 mL/min (1.55 ± 0.1) group was increased and showed reversal after SPK (1.31 ± 0.3, p < 0.001), but not after KTx (1.64 ± 0.1). sTM levels were increased in DN patients and reduced in SPK and KTx recipients (p < 0.05), while the Ang-2/Ang-1 ratio was normalized after SPK and not after KTx alone (from 0.16 ± 0.04 to 0.08 ± 0.02, p < 0.05). Interestingly, in the longitudinal study, reversal of capillary tortuosity and decrease in Ang-2/Ang-1 ratio and sTM was observed within 12 months after SPK. SPK is effective in reversing the systemic microvascular structural abnormalities in DN patients in the first year after transplantation.

Rapid, arteriovenous graft failure due to intimal hyperplasia: a porcine, bilateral, carotid arteriovenous graft model.

BACKGROUND: The loss of patency constitutes the major complication of arteriovenous (AV) polytetrafluoroethylene hemodialysis grafts. In most cases, this graft failure is due to intimal hyperplasia at the venous outflow tract, including proliferation of vascular, smooth muscle cells and fibroblasts with deposition of extracellular matrix proteins. Thus far, procedures developed for improving patency have proven unsuccessful, which can be partly explained by the lack of relevant animal models. For this purpose, we developed a porcine model for AV graft failure that will allow the assessment of promising therapeutic strategies in the near future. MATERIALS AND METHODS: In 14 pigs, AV grafts were created bilaterally between the carotid artery and the jugular vein using expanded polytetrafluoroethylene. Two, 4 or 8 weeks after AV shunting, the grafts and adjacent vessels were excised and underwent histologic analysis. RESULTS: From 2 weeks onwards, a thick neo-intima developed at the venous anastomosis, predominantly consisting of alpha-actin-positive vascular smooth muscle cells (VSMC). Intimal area increased over time, coinciding with a decreased graft flow. Grafts remained patent for at least 4 weeks. At 8 weeks, patency rates declined to less than 50% due to thrombus formation superimposed on progressive neo-intima formation. CONCLUSIONS: Implantation of an AV graft between the carotid artery and jugular vein in pigs causes a rapid neo-intimal response, accompanied by a loss of patency of 50% at 8 weeks after surgery. This model offers a suitable tool to study local interventions aimed at the improvement of AV graft patency rates.