Abstract Background and Aims Chronic kidney disease (CKD) is a growing health problem in Europe exacerbated by aging and the rise of comorbidities such as diabetes mellitus (DM) and hypertension (HTN). Patients with CKD are often multi-morbid and commonly present with cardiovascular (CV) complications. While CV events including myocardial infarction (MI), stroke, and hospitalization for heart failure (HHF) occur more often in patients with advanced CKD, patients with early-stage CKD are also at increased risk. Patients with CKD also experience an increased risk of CV-related mortality, with many patients dying from CV events prior to kidney failure and renal replacement therapy. CV events also present a significant burden to clinicians and healthcare systems due to the high costs and resource use required to manage and treat them. Despite this multidimensional burden, CKD is currently underdiagnosed in Europe, and CKD screening and treatment are rarely included in public health strategies to improve mortality and health outcomes. This study aims to explore the impact of targeted CKD screening followed by guideline-directed therapy in high-risk populations on CV event occurrence and associated treatment costs across Europe. Method The IMPACT CKD microsimulation model simulates CKD progression, CV events, and comorbidities. CKD status is assigned based on estimated glomerular filtration rate (eGFR) and urine albumin-creatinine ratio levels (UACR), and CKD progression is predicted by an annual eGFR decline rate. The model was used to compare two scenarios in four country populations (Germany, Netherlands, Spain, United Kingdom [UK]): 1) targeted screening for people with DM and/or HTN followed by optimal compliance to guideline-directed therapy and 2) current practice (no screening, underdiagnosis, low treatment rates). Annual targeted screening assumed both eGFR and UACR testing. Treatment initiation for people diagnosed with CKD was based on Kidney Disease Improving Global Outcomes guidelines with a 90% compliance assumed to approximate maximum clinical benefit. Therapies were assumed to have a multiplicative treatment effect on eGFR decline and CV events. Current practice assumed observed diagnosis rates without screening and observed treatment rates in each country. For both scenarios, the model projected incidence of CV events and associated management costs in year 10 (2032) and cumulatively over the 10 simulated years (2023-2032). Results Results compare the targeted screening followed by guideline-directed treatment scenario to continuation of current practices for Germany, Netherlands, Spain, and UK (Table 1). The timely diagnosis of patients with CKD and effective treatment was associated with a reduction of 46.7-53.3%, 38.9-41.6%, and 42.1-50.5% in the number of MI, stroke, and HHF events, respectively in 2032 across the four countries. These decreases in the number of CV events were accompanied by proportional reductions in costs to treat the events. Additionally, this policy approach resulted in declines in the cumulative 10-year CV event development and associated healthcare costs, with MI having the largest reductions (40.6-47.2%), followed by HHF (37.4-43.0%), and stroke (29.8-36.1%) across the four countries. Conclusion This study illustrated that early identification of high-risk patients with CKD coupled with guideline-directed treatment would yield substantial clinical and economic benefits from reducing CV events across all four European countries. Fewer MI, stroke, and HHF events were projected in patients with CKD in both 2032 and cumulatively from 2023 to 2032 due to delayed disease progression following effective interventions. Driven by the reduced clinical burden, lower healthcare costs for CV event management were also predicted. These findings support the need for the adoption of policies for early diagnosis and treatment of CKD to slow disease progression and mitigate the growing CV disease burden among patients with CKD and for health systems. Future studies should explore the costs associated with targeted CKD screening and the guideline-directed pharmacological therapies for CKD.

Abstract Background The prevalence of Hepatitis B virus (HBV) related nephropathy particularly membranous nephropathy follows geographic HBV prevalence patterns. Hepatitis B virus infection occurs throughout the world and is endemic in developing countries such as Africa, Eastern Europe, the Middle East, Central Asia, China, Southeast Asia, the Pacific islands and the Amazon basin of South America. Vertical transmission from mother to child often predominates in endemic areas. Membranous glomerulonephritis is the most common histological renal lesion associated with hepatitis B virus infection. Treatment options for renal patients with chronic HBV infection are complex, being dictated by the overall clinical picture and best conducted by multidisciplinary approach and thorough renal monitoring. Therapeutic guidelines are very limited, mostly case series are available in this setting. Anti-viral treatment is indicated in hepatitis B surface antigen (HBsAg)-positive patients with evidence of active liver disease based on aminotransferases and serum HBV DNA levels of 2000-20 000 IU/ml have been proposed as the threshold to start treatment Aim To present a case of young healthy male with chronic HBV infection with nephrotic syndrome and Membranous Nephropathy who developed reactivation after discontinuation of Tenofovir. Case A 32-year old man presented with generalized edema 5 months ago associated with dipstick proteinuria. The swelling has progressed and now associated with walking difficulty thus sought consult and was then admitted. His mother had been hypertensive for more than a decade and work up revealed that she is positive for anti-HBS and negative for HBsAg. His father is negative for HBsAg. His childhood and past medical history is unremarkable. He is a non-smoker, non alcoholic and no history of illicit drug use. He is monogamous and not promiscuous. He has 6 siblings and two of his brothers are hypertensives and found to have chronic Hepatitis B infection. His complement levels were low, ANA and ASO titers were negative. His creatinine was 1.06 mg/dl, BUN- 11.77 mg/dl, SGPT-161 U/L (NV: < 42 U/L), albumin- 1.50 g/dl (NV: 3.5-5.0 g/dl). Total cholesterol- 443.30 mg/dl, Triglycerides 330.46 mg/dl. Urine PCR-4,369 mg/g. Hepatitis profile revealed reactive for HBsAg and HBeAg. His HBV viral load was 170 000 000 IU/ml (989 400 000 copies/ml). His HIV screening was negative. A kidney biopsy was performed which revealed Membranous Glomerulonephritis. He was given Tenofovir 300 mg daily for 6 months. His repeat HBV viral load was < 20 IU/ml (< 116 copies/ml) and his 24 h urinary protein was 1,891 mg/24H. His albumin was 4.57 g/dl and his creatinine remained stable at 0.96 mg/dl while his SGPT went back to normal at 40 U/L. A repeat HBeAg revealed 0.55 which could be a possible seroconversion. Tenofovir was discontinued for 6 months however patient came back with persistent proteinuria associated with high HBV DNA titer. A repeat 24H-urine protein showed 1,182 mg/24H. Tenofovir was resumed and Enalapril was continued. Follow up check up 3 months after revealed urine PCR was 850 mg/g and his kidney function remained stable. He is advised to continue Tenofovir with frequent follow up until he becomes HBsAg-negative. Recommendation We recommend that antiviral treatment in this subset of population requires prolong treatment until persistent seroconversion of HBeAg and persistent undetectable HBV DNA level or have lost HBsAg even if the patient does not develop hepatitis B surface antibody. This recommendation of prolong treatment are still developing and requires further investigation.

Abstract Background and Aims For older people with kidney failure, especially those with comorbidities or poor performance status, the survival benefits of dialysis are uncertain and its quality of life impact greatest. Conservative kidney management (CKM) can be a beneficial alternative. However, there is significant variation in treatment rates among older patients with kidney failure; the CKMAPPS study [1] found rates of older people receiving dialysis ranged from 5-95% across UK renal units. How clinicians communicate about treatment options influences patients’ decision-making, but this has been under-researched. The OSCAR study (Optimising Staff-Patient Communication in Advanced Renal disease) aimed to describe how kidney failure treatment options are presented by renal clinicians to older people (age 65+) with advanced chronic kidney disease (eGFR ≤20 mls/min/1.73 m2 within the last 6 months) and the implications of this for patient engagement with the decision. Method Outpatient consultations between doctors/nurses and eligible patients were recorded at 4 UK renal units with varying rates of CKM. Recorded consultations were screened to identify cases where clinicians presented treatment options for advanced kidney disease. Consultations where clinicians presented both dialysis and CKM were transcribed and analysed in detail using Conversation Analysis, focused on how dialysis and CKM were framed. Post-consultation, patients completed the Shared Decision-Making Questionnaire (SDM-Q-9). Comparisons were made between groups according to how treatment options were presented, using a non-parametric Median Test. Results A total of 110 outpatient consultations were recorded (104 audiovisual, 6 audio). Recordings included 38 doctors and nurses and 94 patients; mean patient age 77 (65-97), 33 female/61 male, mean eGFR 15 (range 4-23). Sequences where clinicians presented both dialysis and CKM as treatment options were analysed (n = 21). Two approaches to presenting CKM were identified: 1) CKM as a main option (n = 6; see Fig. 1), 2) CKM as a subordinate option (n = 15; see Fig. 2). The mean consultation length was the same in both groups (23 mins). Recurrent features of the first approach included: framing CKM as having potential personal benefits to the patient; explicitly labelling it as a treatment option; not framing it as an option preferred by or relevant to only a minority of patients. In contrast, when CKM was presented as a subordinate option, recurrent features included: framing CKM as not having benefit to the patient; not explicitly labelling CKM as a treatment option; appending CKM to the main decision-making sequence; framing CKM as an option only chosen by a minority of patients. Presenting CKM as a main option alongside dialysis was a less common approach (n = 6 vs. n = 15), but associated with more interactional opportunities for patients to ask questions about CKM, assert their perspective, and assess CKM as a relevant option, as well as significantly higher patient ratings of shared decision-making (total SDM-Q-9 score, p = 0.041). Conclusion This is the first fine-grained analysis of the relationship between the conversational practices used by clinicians and their impact on patient engagement with treatment options and ratings of shared decision-making. Despite evidence that dialysis does not reliably extend older patients’ lives at acceptable costs to quality of life, we found that clinicians tend to present dialysis as the default treatment and CKM as subordinate, if at all. Our findings demonstrate that presenting treatment options is not enough; how clinicians present options has important implications for patient engagement in shared decision-making. To provide patients with the opportunity to evaluate CKM as a valid option requires clinicians to clearly detail the advantages and disadvantages of both treatments. Study findings will form the basis of a new communication training intervention for clinicians.

Abstract Background and Aims The diversity in blood pressure (BP) response to various salt intakes, known as Salt Sensitivity (SS), exhibits significant differences among individuals within the hypertensive subjects relative to the general population. Hypertension (HT) and SS are correlated to elevated levels of plasma Endogenous Ouabain (EO). We aimed at delineating the role of the missense variation rs2254524 (Val642Leu; C>A) within the Lanosterol Synthase gene (LSS), encoding for an enzyme involved in steroid biosynthesis. Individuals with the AA LSS variant, adherent to a low-salt diet, had a more significant BP reduction compared to those with CC genotype. Method Utilising CRISPR-Cas9, we established a knock-in mouse model carrying the analogous LssV643L/V643L. Male mice were subjected to a normal-salt diet (0.5% NaCl), high-salt diet (4% NaCl), or low-salt diet (<0.03% Na) and BP was monitored every two days using a tail-cuff system for 10 days. Blood and urine were collected separately in metabolic cages. Results Homozygous mice for Lss mutation demonstrated normal viability and healthy phenotype, undistinguishable from the WT. At baseline, the Lss AA mice were affected by a significant augmentation in kidney weight at 3 and 12 months of age, relative to WT. The Lss V643L does not influence EO levels or systolic BP (SBP), specifically at 3 and 12 months, but affects the SBP response to dietary salt. AA mice showed a high SBP in a high-salt diet against control diet at the same time intervals. Moreover, the Lss variant impacted the pressure-natriuresis relationship, with the mutated AA strain showing a less steep curve both in control diet versus the high-salt diet and in control diet versus the low-salt diet, displaying the SS phenotype of the AA mice. The 12-month-old mice in high-salt diet displayed cardiac hypertrophy with the AA genotype exhibiting an elevated prevalence of cardiac fibrosis. The level of Lss mRNA decreased in the adrenal glands in response to a high-salt diet. RNASeq analysis identified distinct patterns in several Slc genes, Cbr2 and Arhgap26 within the kidneys of these mice, in control or high-salt dietary conditions. Conclusion The established LssV643L/V643L mouse model effectively mirrors the SS-HT phenotype. Our findings highlight the significance of the Lss in modulating BP in response to salt stimuli suggesting a potential organ damage pathway requiring further exploration in older mice.

Abstract Background and Aims Despite the demonstrated efficacy of a low-protein diet (LPD) in slowing the progression of chronic kidney disease (CKD), the underlying mechanisms remain inadequately understood. Given the potential drawbacks, including poor compliance and a risk of malnutrition, gaining insights into the molecular mechanisms driving these benefits is crucial for safe clinical translation. In this study, our objectives were to 1) assess the impact of an LPD on the metabolic health of individuals and mice with CKD; 2) investigate the hypothesis that these effects are mediated by a reduction in uremic toxins; and 3) explore whether the combination of an LPD and supplementation with a strain of Lactiplantibacillus plantarum (strain LpWJL), selected for its ability to enhance growth in a Drosophila model of diet-induced stunting, could synergistically amplify the positive effects of the LPD while minimizing its potential adverse consequences. Method We recruited CKD patients with no diabetes or prediabetes, who were randomized to a LPD and cetoanalogues (0.4 g/kg/day) (n = 5) or a normal diet (ND) (0.8 g/kg/day, n = 6) for 3 months. A glucose tolerance test was measured at baseline and after 3 months. Four groups of mice were studied: a sham group with a ND (n = 4), 5/6th nephrectomized mice with a ND (n = 7), and 5/6th nephrectomized mice with LPD (5% w/w) and LPwjl (n = 7) or placebo (n = 7) for 6 weeks. Blood samples and liver tissues were analyzed with the MxP 500 Quant® (Biocrates) kit by LC-MS/MS (XEVO TQ-XS, Waters). Results Under LPD, beta cell function showed improvement (Matsuda index, P = 0.049, and AUC insulin, P = 0.02). Although the diet did not influence body composition, the LPD group exhibited a reduction in muscle strength (P = 0.02). LPD decreased blood concentration several uremic toxins such as indoxyl sulfate and TMAO. Insulin sensitivity and beta-cell function demonstrated negative correlations with urea, TMAO, and indoxyl sulfate. Mouse studies confirmed the positive impact of LPD on glucose homeostasis but also revealed detrimental effects on lean mass and fat mass. Supplementation with LpWJL during LPD mitigated the loss of fat mass, improved the insulin tolerance test, and enhanced the reduction of certain blood uremic toxins, including indoxyl sulfate. LpWJL supplementation exerted a substantial impact on the hepatic metabolome, resulting in increased triglyceride accumulation and decreased levels of diglycerides, acylcarnitines, and phosphocholines. Conclusion We have highlighted the substantial benefits of a 3-month LPD on markers of glucose homeostasis and uremic toxins in mice and humans without diabetes. LpWJL induced profound modifications in both blood and liver metabolome and was associated with a limitation of trophic alterations in LPD diet-induced CKD mice, while also enhancing glucose homeostasis. Further studies are essential to gain a better understanding of the underlying mechanisms involved.

Abstract Background and Aim Renal involvement of systemic lupus erythematosus (SLE) is known as lupus nephritis (LN). The prognosis of lupus patients with LN is worse than those without kidney involvement. Renal survival can be improved in patients with lupus nephritis with appropriate and timely treatment. The aim of our study is to compare the clinical and pathological findings of patients diagnosed with LN in our clinic by renal biopsy, the results of the treatment applied in our center, and renal survival with the literature. Method The local ethical committee approved the study design (date: 04/10/2021; reference number: 121/07) and all procedures were conducted in accordance with the Declaration of Helsinki. Forty-four patients diagnosed with lupus nephritis between January 2012 and January 2021 in the Nephrology Department of Ankara Dıskapı Yıldırım Beyazıt Education and Research Hospital were enrolled in this study. Patient data were analyzed retrospectively. Exclusion criteria were as follows: 1) Patients under 18 years of age, 2) patients with follow-up of period less than 6 months and who did not attend regular follow-ups, 3) patients with connective tissue diseases other than SLE. Results The mean age of the study population was 36.0 ± 10.8, and the majority were female. All patients had proteinuria at diagnosis. Class IV LN was the most common (36.4%). Majority of the patients were in the proliferative LN group (65.9%). A significant difference was found between patients diagnosed with proliferative LN (n = 29) and non-proliferative LN (n = 15) in terms of high serum creatinine levels, low serum complement, presence of anti-dsDNA antibodies and active urinary sediment with proliferative LN (p = 0.021, p = 0.024, p = 0.008, p = 0.008). Treatment responses at 6 and 12 months are shown in Table 1. A significant difference was found between the patients who were in complete remission (n = 16) at 12 months and those who were not in remission (n = 27) in terms of low systolic blood pressure and the estimated glomerular filtration rate (eGFR) at the time of diagnosis (p = 0.008, p = 0.016). It was observed that most of the patients who were not in complete remission at 12 months were in the proliferative LN group (p = 0.024). There was no significant difference in the partial and complete remission rates at 6 and 12 months between the group receiving steroids + cyclophosphamide (n = 14) and the group receiving steroid + mycophenolate mofetil (MMF) (n = 15) in the initial treatment of proliferative LN (p>0.05). In the group taking MMF treatment (n = 15) had a shorter time to reach complete remission than the group taking cyclophosphamide treatment, and it was statistically significant (Table 2) (p = 0.048). It was found that end-stage renal disease (ESRD) developed in 9% of the patients and 2 patients died during followed up period. Recurrence was detected in 8 patients, and it was observed that more than half of the patients with relapse could not achieve complete remission at 12 months. Patients with relapse had higher systolic blood pressure and serum total cholesterol level at the time of diagnosis compared to patients without recurrence (p = 0.013, p = 0.045). In addition, patients with relapse were found to have a higher level of proteinuria at the time of diagnosis (p = 0.036). Conclusion Lupus nephritis remains an important source of morbidity and mortality for SLE patients. SLE patients should be regularly examined for renal involvement. About 10-20% of patients diagnosed with lupus nephritis develop ESRD. Early diagnosis of LN and initiating appropriate treatment when diagnosed without delay is important for renal and patient survival. Although there were some different results, most of the statistically significant data in our study were found to be compatible with the existing literature. However, these data need to be confirmed by studies with high patient participation.

Abstract Background and Aims Refractory lupus nephritis (LN) is defined as either no response to standard of care (SOC), i.e., failure to improve within 3–4 months or not achieving partial response within 12 months or complete response after 2 years of treatment. This condition is characterized by poor outcome, is often life-threatening, and represents a relatively unexplored clinical setting. A dysregulation of CD38 expression has been reported in Systemic lupus Erythematosus, and highly expressing CD38 plasma cells relegated in bone marrow niches inaccessible to conventional and biological agents could represent a source of repowering of the immune dysregulation. The efficacy and safety of daratumumab, a monoclonal antibody specifically directed at CD38, given without any other immunosuppressant or agents targeting B-cell-activating factor, were assessed in a discrete cohort of patients with refractory LN, in whom both SOC and rescue treatments had failed. Method Six patients (1 male and 5 females) aged 41.3 years (range 20 to 61 years) were treated with Daratumumab monotherapy. The treatment protocol consisted of 16 mg/kg daratumumab administered intravenously weekly for 8 weeks, then every two weeks 8 more times, and lastly monthly for 8 times. All patients had failed previous treatments with both SOC including either mycophenolate mofetil or cyclophosphamide-azathioprine, and rescue therapies including Rituximab, Ocrelizumab, Belimumab, and iv IgG. Results One out of six patients did not show clinical response after 6 months of therapy, and Daratumumab was discontinued. Five patients continued to be treated (total treatment maximum 22 infusions) and reached a 12-month observation. Renal biopsy performed before daratumumab administration revealed a class IV LN in 1 patient, class V LN in 1 patient, class III+V LN in 1 patient and class IV+V LN in the other 2. Three patients achieved a complete renal response and the other two a partial renal response. A significant decrease in proteinuria from 5.6 g per 24 h to 0.59 g per 24 h, 0.9 g per 24 h and 0.8 g per 24 h (P = 0.0010) at 3 months, 6 months and 12 months, respectively, was observed. The mean value of serum Creatinine (sCr) decreased from 2.3 to 1.5 mg/dl. The mean value of sCr decreased from 2.3 mg dl−1 to 1.5 mg/dl (P = 0.98) at 12 months. C3 and C4 levels increased from 72.8 (range, 0–99) to 101.6 (range, 78.0–135) mg/dl (P = 0.16) and from 9.4 (range, 5–14) to 20.4 (range, 13–30) mg/dl (P = 0.0182), respectively. A decrease in IgG mean levels from 971.7 mg dl−1 (range, 251–1546) to 512.3 mg/dl (range, 88–957), 481.7 (range, 84–878) mg/dl and 508.4 mg/dl (range, 192–685) at 3 months, 6 months and 12 months, respectively, was observed. Improvement of clinical symptoms was paralleled by seroconversion of anti-double-stranded DNA antibodies (p = 0.03), significant decrease in interferon-gamma values (p = 0.0006), BMCA-B-cell maturation antigen (p = 0.0005) and soluble CD163 levels (p = 0.045), and IL 10 levels (p = 0.0006). Clinical remission was substantiated by improvement of SLEDAI-2K score (p = 0.03). Daratumumab was generally well tolerated. Conclusion These data suggest that Daratumumab administered alone (i.e., without any other immunosuppressant or agents targeting B-cell activating factor) is highly effective in refractory LN. The multifaced effects of Daratumumab on the inflammatory burden and the interferon gamma profile could provide a restoration of the immune balance.

Abstract Background and Aims Subpopulations of CD4+ cells, play a major role in mediation of rejection and tolerance to a renal transplant. This study examined subsets of CD4+ T cells and CD4+CD25+CD127lo Treg in blood of renal graft recipients with grafts surviving >10 years for changes in lymphocyte subpopulations that may indicate transplant tolerance and potentially identify patients who could reduce immunosuppression. Based on CD45RA and CD25/Foxp3 expression, Miyara et al identified five populations (Pop) within CD4+ cells. Pop I, II and III are T regulatory cells (Treg); Pop I as naïve (CD45RA+Foxp3+) Treg, Pop II as activated (CD45RA-Foxp3hi) Treg, and Pop III as cytokine secreting (CD45RA-Foxp3+) cells which also includes activated effector cells. Pop IV are activated T cells (CD45RA-Foxp3−), and Pop V are naïve T cells (CD45RA+Foxp3−). Here, we examined three Treg populations (Pop I-III) within CD4+CD25+CD127lo Treg, and Pop IV and V within CD4+ T cells. To further characterise these subpopulations, we examined chemokine receptor expression (CXCR3, CCR4, CCR6, and CCR7) to identify T helper phenotype (Th-like) of Treg populations. Th1-like Treg were identified as CCR4+CXCR3+CCR6−, Th2-like as CCR4+CXCR3-CCR6−, Th17-like as CCR4+CXCR3-CCR6+, and Th1/Th17-like as CCR4+CXCR3+CCR6+. We also examined expression of activated Treg effector molecules CD39, Class II MHC, and PD-1. Method Peripheral blood mononuclear cells (PBMC) were isolated from fresh blood collected from healthy volunteers (HV) (n = 44) and long-term stable renal transplant patients with grafts surviving >10 yrs (RT) (n = 25). Panels of monoclonal antibodies for Treg (CD4/CD25/CD127/CD45RA/Foxp3), chemokine receptors (CXCR3, CCR4, CCR6, CCR7), and Treg activation and suppression-related molecules (CD39/HLA-DR/PD-1) were also used. Data was acquired on BD FACSCanto II using BD FACSDiva software (v8.0) and analysed using FlowJo. Lymphocyte populations were examined after FSC vs SSC gating and doublets exclusion. T cell and Treg populations were analysed within CD4+ and CD4+CD25+CD127lo T cells. Results RT had lower lymphocyte counts than HV (p = 0.0156) but had a similar percentage of CD4+ T cells. Treg were significantly lower in RT compared to HV (p = 0.0012). RT patients had less naïve Treg (Pop I, p = 0.0083) and naïve effector T cells (Pop V, p = 0.0003), but had higher activated effector T cells (Pop IV, p = 0.0002) than HV. There were no differences in activated Treg Pop II or III between the two groups. There were significantly less cells expressing CXCR3 in RT than HV in Pop I (p = 0.0120), Pop II (p < 0.0001), Pop III (p < 0.0001) and Pop IV (p = 0.0047). Similarly, RT had fewer cells expressing CCR6 compared to HV in Pop II (p = 0.0019), Pop III (p = 0.0015), and Pop V (p = 0.0017). RT had lower proportions of CCR7 expressing cells in Pop I (p = 0.0035), Pop IV (p = 0.0097), and Pop V (p = 0.0002), but higher in Pop II (p = 0.0041) compared to HV. There were similar percentages of cells with Th1 and Th17-like phenotypes in both groups. However, RT had significantly more Th2-like Treg in Pop II (p = 0.0033) and Pop III (p = 0.0100), and significantly less Th1/Th17-like Treg in Pop II (p = 0.0005) and Pop III (p = 0.0017) compared to HV. There were little to no differences in the Treg activation and suppression-related molecules within subpopulations of Treg. Conclusion Long-surviving RT patients had less naïve Treg and naïve effector cells, but increased proportions of activated effector cells. There were little to no differences in the expression of Treg activation or suppression-related molecules in Treg subpopulations. Unexpectedly, activated Treg in RT had significantly higher proportions with a Th2 phenotype and significantly less Treg with a Th1/Th17 phenotype. Whether this is a consequence of immunosuppression, or a biological shift associated with tolerance, remains to be resolved. Further studies are required to investigate the relevance of these changes in identifying induction of transplant tolerance.