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Hyponatremia is the most common electrolyte disorder and it has been associated with increased mortality.This study evaluated hyponatremia as a prognostic factor for severity and mortality.We compared the prevalence of hyponatremia among patients who died during the year 2017 (from 1 January 2017 to 31 December 2017) with the prevalence of hyponatremia among subgroups of patients, i.e. outpatients, patients hospitalized for more than 2 days and patients admitted in the intensive care unit (ICU). We also described the mortality rate and the prevalence of comorbidities among hyponatremic patients, according to hyponatremia degree (slight, moderate, severe), basal characteristics, comorbidities and their outcome (discharged, hospitalized or died).In our population of a public hospital setting, hyponatremia was present at admission in 17% of deaths, and the comparison between hyponatremic and normonatremic patients in terms of mortality confirms the hypothesis that this disorder is in anyway strictly associated with vulnerability and with a poor prognosis.We conclude that hyponatremia is a predictive marker for a bad clinical course, therefore patients with this electrolyte disorder should be carefully monitored.

Peripheral blood mononuclear leukocytes (PBL) of uremic patients (u-PBL) prematurely die by apoptosis, thus sustaining leukopenia and immune dysfunction. Uremic retention solutes have been alleged to playing a causal role in this immune cell defect. However, both the molecular identity and pro-apoptotic mechanism of these solutes remain poorly characterized. In this study, we prepared a fraction of the uremic plasma (u-Pl) rich in these solutes (proteinaceous material with molecular weight > 50 kDa, namely the uremic-high MW fraction or u-HMW) that was used to demonstrate their pro-apoptotic activity in u-PBL. Such a detrimental activity was also confirmed in THP-1 and K562 mononuclear cells in association with increased cellular generation and secretion of H2O2, and JNK/cJun-dependent apoptotic signaling downstream of the endoplasmic reticulum stress response protein IRE1-α. The u-HMW also induced autophagy in THP-1 mononuclear leukocytes. These alterations of u-PBL proteostasis were associated with the presence in the proteome of these cells, but not of control PBL, of the main proteins and protein decoration targets (assessed by 2,4-diphenylhydrazine derivatization) of u-Pl and thus of u-HMW, namely albumin, transferrin and fibrinogen. These findings demonstrate that large solutes induce apoptosis in u-PBL leading to abnormal plasma protein endocytosis and terminal alteration of cellular proteostasis mechanisms. We define this response of PBL to large uremic solutes as the “immuno-proteostasis response” (IPR) of uremia.

Elderly individuals with chronic disorders tend to develop inflammaging, a condition associated with elevated levels of blood inflammatory markers, and increased susceptibility to chronic disease progression. Native and adaptive immunity are both involved in immune system senescence, kidney fibrosis and aging. The innate immune system is characterized by a limited number of receptors, constantly challenged by self and non-self stimuli. Circulating and kidney resident myeloid and lymphoid cells are all equipped with pattern recognition receptors (PRRs). Recent reports on PRRs show kidney overexpression of toll-like receptors (TLRs) in inflammaging autoimmune renal diseases, vasculitis, acute kidney injury and kidney transplant rejection. TLR upregulation leads to proinflammatory cytokine induction, fibrosis, and chronic kidney disease progression. TLR2 blockade in a murine model of renal ischemia reperfusion injury prevented the escape of natural killer cells and neutrophils by inflammaging kidney injury. Tumor necrosis factor-α blockade in endothelial cells with senescence-associated secretory phenotype significantly reduced interleukin-6 release. These findings should encourage experimental and translational clinical trials aimed at modulating renal inflammaging by native immunity blockade.

We propose a new organ-conditioning strategy based on mesenchymal stromal cell (MSCs)/extracellular vesicle (EVs) delivery during hypothermic perfusion. MSCs/EVs marker CD73 is present on renal proximal tubular cells, and it protects against renal ischemia-reperfusion injury by converting adenosine monophosphate into adenosine (ADO). In this study, after checking if CD73-silenced EVs (EVsi) would impact in vitro tubular-cell proliferation, we perfused kidneys of a rat model of donation after circulatory death, with Belzer solution (BS) alone, BS supplemented with MSCs, EVs, or EVsi. The ADO and ATP levels were measured in the effluents and tissues. Global renal ischemic damage score (GRS), and tubular cell proliferation index (IPT) were evaluated in the tissue. EVsi did not induce cell proliferation in vitro. Ex vivo kidneys perfused with BS or BS + EVsi showed the worst GRS and higher effluent ADO levels than the MSC- and EV-perfused kidneys. In the EV-perfused kidneys, the tissue and effluent ATP levels and IPT were the highest, but not if CD73 was silenced. Tissue ATP content was positively correlated with tissue ADO content and negatively correlated with effluent ADO level in all groups. In conclusion, kidney conditioning with EVs protects against ischemic damage by activating the CD73/ADO system.

Patients with severe COVID-19 both seroconvert earlier and develop higher concentrations of SARS- CoV-2-specific IgG than patients with mild symptoms. In this retrospective study we considered different categories of patients defined as "vulnerable" because affected by other pathologies, such as patients with genetic and cardiovascular diseases; patients with autoimmune dermatological dis- ease; kidney and lung transplant patients, and pregnant women because the prevalence of Covid-19 infection during pregnancy is not known. This study was performed at IRCCS San Matteo Hospital in Pavia, North Italy, a zone considered at high risk during the COVID-19 pandemic from June to December 2020. None of the positive screened patients had symptoms of COVID-19 infection at the time of inclusion in this study.

Post-transplant neutropenia (PTN) is frequently reported in the first-year after transplantation. Although prevalence and clinical consequences are widely described, there are no guidelines to manage diagnosis and treatment. We report here a case of persistent PTN occurred in a patient undergoing a kidney transplant from an AB0-incompatible living donor. The desensitization protocol consisted of Rituximab administration and immunoadsorption while the pre-transplant protocol, which was initiated 14 days before the transplant, included Tacrolimus, Mofetil Mycophenolate (MMF), antimicrobial and antiviral prophylaxis. Induction therapy consisted of anti-thymocyte globulins and steroids, while maintenance after transplantation consisted of steroid, tacrolimus and MMF. When the first occurrence of leukopenia was observed six weeks after the transplant, firstly antimicrobial/antiviral prophylaxis was stopped and later also MMF treatment was interrupted but severe neutropenia relapsed after MMF resuming treatment. Immunological and virological causes were excluded. The patient was treated with Filgrastim. Bone marrow biopsy, which was performed to exclude a hematological cause of severe persistent neutropenia, revealed a bone marrow hypoplasia with neutrophils maturation interrupted at the early stages. This case highlights the need to establish diagnostic and therapeutic guidelines for PTN which take in consideration all the therapeutic steps including the pre-transplant phase in particular in the context of AB0i where immunosuppression is more consistent.

We report a preliminary experience of adjuvant therapy with Hemoperfusion (HP) in patients with Severe Acute Respiratory Syndrome-CoronaVirus 2 (SARS-CoV2) pneumonia. Currently, there are no approved treatments for CoronaVirus Disease 19 (COVID-19); however, therapeutic strategies based on the preclinical evidence include supportive measures, such as oxygen supplementation, antiviral, and anticoagulant agents. Despite these treatments, 10% of patients worsen and develop severe acute respiratory distress syndrome (ARDS). Since the pathogenic mechanism of ARDS is an uncontrolled inflammatory state, we speculate that removing inflammation effectors from blood may contrast tissue injury and improve clinical outcome. In a scenario of dramatic medical emergency, we conducted an observational study on 9 consecutive patients hospitalized in COVID Intensive Care Unit, where 5 of 9 consecutive patients were treated with HP, due to the emergency overload made it impossible to deliver blood purification in the other 4 patients. COVID-19 was diagnosed through the identification of virus sequences by reverse transcription-PCR on respiratory specimens. All patients had severe pneumonia requiring continuous positive airway pressure. HP was started in all patients 6–7 days after hospital admission. The treated patients (T) received 2 consecutive sessions of HP using CytoSorb cartridge. Our results show a better clinical course of T compared to control patients (C), in fact all T except 1 survived, and only 2 of them were intubated, while all C required intubation and died. Lymphocytopenia worsened in C but not in T. C-reactive protein decreased in both patients, but to a greater extent in T. IL-6, IL-8, and TNF-α decreased after HP, IL-10 did not change. Respiratory function remained stable and did not worsen in T compared to C. The limited sample size and observational study design preclude a sound statement about the potential effectiveness of HP in COVID-19 patients, but our experience suggests a potential therapeutic role of adjuvant CytoSorb HP in the early course of CO­VID-19 pneumonia. A randomized clinical trial is ongoing.

Introduction: Acute kidney injury (AKI) is a common complication among hospitalized patients, potentially affecting short- and long-term clinical outcomes. In this retrospective study, we evaluated renal outcomes in noncritically ill patients who required acute hemodialysis (HD) because of an AKI episode occurring during hospitalization. Methods: Sixty-three hemodynamically stable patients with AKI undergoing acute intermittent HD were included. Kidney function was evaluated at baseline control (pre-AKI), at AKI diagnosis and during the follow-up. According to serum creatinine and the estimated glomerular filtration rate (eGFR), we defined three clinical conditions: renal recovery, different stages of acute kidney disease (AKD), and chronic kidney disease (CKD). Results: Among the 63 patients evaluated, 34 patients (54%) had a history of CKD. Six patients (10%) presented early full renal recovery. HD treatment was stopped in 38 patients (60%), while 25 patients (40%) required maintenance HD. Dialysis-independent patients presented lower comorbidity and higher baseline eGFR and delta creatinine, compared to dialysis-dependent patients. Baseline CKD, previous AKI episodes, and parenchymal causes of AKI were associated with a significant risk of dialysis dependence. At 1-month control, 15 patients (39%) presented AKD stage 0, 6 patients (16%) AKD stage 1, and 17 patients (44%) AKD stage 2–3. At 3-month control, 29 out of 38 patients recovering from AKI (76%) presented CKD. AKD stage was significantly correlated with the risk of CKD development, which, resulted higher in patients with lower baseline eGFR. Conclusions: AKI might represent a risk factor for the development of chronic kidney damage, even in noncritically ill patients.

Coronavirus is a high-risk pathogen for kidney transplant recipients receiving immunosuppressive therapy; Coronavirus disease 2019 (COVID-19) is an infection causing severe acute respiratory syndrome (SARS-CoV2), and progressive withdrawal of immunosuppressive drugs has been suggested in transplanted patients. At IRCCS San Matteo Hospital in Pavia, Northern Italy, during the pandemic we performed a screening and all transplanted patients were evaluated for IgG anti SARS-CoV-2; 12 of 201 kidney transplant recipients (6%) screened for IgG anti SARS-CoV-2 (s) developed kidney transplant rejection; 10 (83%) were negative and 2 (17%) resulted positive for SARS-CoV-2 IgG, while among 189 patients without rejection, 162 (86%) resulted negative and 27 (14%) positive (P=0.69). COVID 19 course may be more severe in kidney transplant recipients but it does not significantly increase risk of kidney rejection.

Donation after circulatory death (DCD) allows expansion of the donor pool. We report on 11 years of Italian experience by comparing the outcome of grafts from DCD and extracorporeal membrane oxygenation (ECMO) prior to death donation (EPD), a new donor category. We studied 58 kidney recipients from DCD or EPD and collected donor/recipient clinical characteristics. Primary non function (PNF) and delayed graft function (DGF) rates, dialysis need, hospitalization duration, and patient and graft survival rates were compared. The estimated glomerular filtration rate (eGFR) was measured throughout the follow-up. Better clinical outcomes were achieved with EPD than with DCD despite similar graft and patient survival rates The total warm ischemia time (WIT) was longer in the DCD group than in the EPD group. Pure WIT was the highest in the class II group. The DGF rate was higher in the DCD group than in the EPD group. PNF rate was similar in the groups. Dialysis need was the greatest and hospitalization the longest in the class II DCD group. eGFR was lower in the class II DCD group than in the EPD group. Our results indicate good clinical outcomes of kidney transplants from DCD despite the long “no-touch period” and show that ECMO in the procurement phase improves graft outcome, suggesting EPD as a source for pool expansion.

Simple Summary The most common cause of late allograft failure is chronic active antibody-mediated rejection (ABMR), but no effective therapy is available. Different immunosuppressive drugs in combination with procedures that remove serum antibodies have been used and the results have not shown to improve graft and patient outcome, but only an increased risk of adverse events. Extracorporeal pho-topheresis (ECP) is leukapheresis-based immunomodulatory therapy not associated with adverse effect, in which lymphocytes treat-ed with 8-methoxypsoralen (8-MOP) are irradiated with ultraviolet-A (UVA) ex vivo and re-infused into the patient. In this study we investigated therapeutic long-term effect of ECP in patients with biopsy proved chronic ABMR. Abstract Objective: Chronic renal antibody-mediated rejection (ABMR) is a common cause of allograft failure, but an effective therapy is not available. Extracorporeal photopheresis (ECP) has been proven successful in chronic lung and heart rejection, and graft versus host disease. The aim of this study was to evaluate the effectiveness of ECP in chronic ABMR patients. Patients and Methods: We investigated ECP treatment in 14 patients with biopsy-proven chronic ABMR and stage 2–3 chronic renal failure. The primary aim was to e valuate the eGFR lowering after 1 year of ECP therapy. The ECP responders (R) showed eGFR reduction greater than 20% vs the basal levels. We also evaluated the effectiveness of ECP on proteinuria, anti-HLA antibodies (HLAab), interleukin 6 (IL-6) serum levels, and CD3, CD4, CD8, CD19, NK, Treg and T helper 17 (Th17) circulating cells. Results: Three patients dropped out of the study. The R patients were eight (72.7%) out of the 11 remaining patients. Because ECP was not associated with any adverse reaction, the R patients continued such treatment for up to 3 years, showing a persisting eGFR stabilization. Twenty four hour proteinuria did not increase in the R patients over the follow-up when compared to the non-responder patients (NR). In the R patients, the HLAab levels were reduced and completely cleared in six out of eight patients when compared with the NR patients. The NR HLAab levels also increased after the discontinuation of the ECP. The ECP in the R patients showed a decrease in CD3, CD4, CD8, CD19, and NK circulating cells. The ECP treatment in the R patients also induced Tregs and Th17 cell increases, and a decrease of the IL-6 serum levels. Conclusions: ECP abates the HLAab titer and renal failure progression in patients with chronic renal ABMR, modulating the immune cellular and humoral responses.

Background and Aims Chronic antibody-mediated rejection (cABMR) is the main cause of kidney allograft failure. Currently there is no effective treatment. Extracorporeal photopheresis (ECP) is an immunomodulating therapy that acts increasing regulatory T cells and reducing effector T cells. ECP has been used with success to treat heart and lung rejection, acute and chronic Graft versus Host Disease and few cases of acute antibody mediated kidney rejection. The aim of this study was to describe the preliminary experience of ECP use to treat kidney cABMR. Method This is an ongoing prospective observational study approved by the ethical committee of our institution. The study started in 2017 on kidney transplant recipients (KTR) with cABMR diagnosis bioptically proved. All patients received ECP in addition to the standard treatment. ECP schedule is reported in Figure 1. Serum and urine samples were collected at the beginning of the study and every 6 months during the follow up period. Analyzed variables were: glomerular filtration rate (GFR), urine albumin-creatinine ratio (UACR), Donor Specific Antibodies (DSA) measeured as Mean Intensity Fluorescence (MIF) by Luminex. Collected clinical data included: death, hospital admission number, cancer diagnosis, infections. Quality of life (QoL) scale was also recorded. Results We enrolled 6 KTR equally distributed in gender (3 M, 3 F) and aged 49.16 ± 7.15 years (mean±ds) . Median follow up time was 12 months (min 12 – max 36). All KTR had both class I and class II DSA with MFI > 5000 (min 5000 – max 36780). In all patients no GFR deterioration was observed during follow up. UACR improved in 3 KTR while remained stable in the other 3 patients. No deaths or infections or cancer diagnosis or hospital admissions occurred. QoL ameliorated in all KTR. DSA MIF decreased in all KTR and in 3 of them became even negative after 12 months of treatment. Conclusion ECP is a safe and effective treatment to slow down the deterioration of kidney function in cABMR patients. Further studies are necessary to identify the responder patients and to better tailor the treatment schedule.

Background and Aims Adenosine (Ado), the main substrate of ATP, is a potent endogenous anti-inflammatory nucleoside. Hypoxia induces ATP depletion, AMP extracellular increase that is phosphohydrolyzed to ADO by the enzyme ecto-5′-nucleotidase (CD73). Constitutive CD73 expression is higher in deep cortex outer medulla that is the most vulnerable region to ischemic injury. Notably CD73 is also expressed on Mesenchymal Stromal Cell (MSC) and is essential phenotypic marker. Previously in a rat model of Donation after Circulatory Death (DCD), we demonstrated that the perfusion of ischemic kidney with MSC or MSC derived Extracellular Vesicles (EV) protects tissue up regulating mitochondrial energetic metabolism genes. The aim of this study was to investigate the role of CD73/Ado system in MSC/EV protection from ischemia. Method Fisher rats were used as kidney donors and Lewis rats as MSC donors. MSC missing CD73 were produced by electroporation in the presence of specific siRNA to block CD73 expression (siMSC). EV were isolated from supernatants of MSC and siMSC (siEV) medium through differential ultracentrifugations. Size distribution and enumeration analysis were performed using NanoSight NS300. EV phenotypic characterization was performed in flow cytometer using CD45, CD49e, CD63, CD9, CD81 and CD73 monoclonal antibodies. DCD model was obtained by renal artery clamping for 20 minutes. This warm ischemia time represents the “no touch period” imposed by the Italian law to death declaration. After nephrectomy, kidneys were perfused in hypothermia (4°C) with Belzer Solution (BS), or BS supplemented with 3x106 MSC (MSC) or BS supplemented with 28,5x109 EV/siEV (EV/siEV). The effluent fluid (EF) was collected at the beginning (T0), every hours (T1, T2, T3) and at the end of the perfusion (T4). Ado and ATP determinations in EF and tissues were performed by HPLC and ELISA, respectively. Results EF Ado was significantly higher in MSC vs BS from T1 and in EV vs BS from T0 to T4 (p Conclusion CD73 expressed on MSC /EV impacts on cell energy metabolism pathway and ATP generation. This is the first evidence that MSC/EV act through CD73/Ado system to prevent ischaemic damage.

Background. In this study we investigated the relevance of myostatin and Hepatocyte Growth Factor (HGF) in patients undergoing hemodialysis HD and the influence of different HD modalities on their levels. Methods. We performed a prospective crossover study in which HD patients were randomized to undergo 3-month treatment periods with bicarbonate hemodialysis (BHD) followed by online hemodiafiltration (HDF). Clinical data, laboratory parameters, and myostatin and HGF serum levels were collected and compared. Results. Ten patients and six controls (C) were evaluated. In any experimental condition myostatin and HGF levels were higher in HD than in C. At enrollment and after BHD there were not significant correlations, whereas at the end of the HDF treatment period myostatin and HGF were inversely correlated (r -0.65, p<0.05), myostatin serum levels inversely correlated with transferrin (r -0.73, p<0.05), and HGF levels that resulted positively correlated with BMI (r 0.67, p<0.05). Moving from BHD to HDF, clinical and laboratory parameters were unchanged, as well as serum HGF, whereas myostatin levels significantly decreased (6.3 ± 4.1 versus 4.3 ± 3.1 ng/ml, p<0.05). Conclusions. Modulation of myostatin levels and myostatin/HGF balance by the use of different HD modalities might represent a novel approach to the prevention and treatment of HD-related muscle wasting syndrome.

The role of vitamin E in both enzymatic and free radical-dependent metabolism of polyunsaturated fatty acids (PUFAs) has been well demonstrated. This study proposed a new LC-MS/MS method to quantify the main vitamin E forms, their metabolites and main PUFA species in human blood, since, at present, there are not procedures able to simultaneously determine these two classes of compounds. After the optimization of sample treatment and reverse-phase separation conditions, tandem mass spectrometry detection was evaluated experimenting both positive and negative electrospray ionisation modes. The procedure was also preliminarily adapted to assess five arachidonic acid-derived eicosanoids that could be under the influence of vitamin E function, such as LTB4 (leukotriene B4), 20-HETE (20-hydroxyeicosatetraenoic acid) and their ω-oxidation metabolites. After the validation study, the performance characteristics were confirmed analysing a certified reference material (SRM® 1950 - frozen human plasma by NIST). Finally, an application of the method in the analysis of lipid abnormalities of chronic kidney disease patients was shown.

Aim To evaluate the effects on glycemic excursions during bicarbonate dialysis (BHD) compared to hemodiafiltration (HDF) in type 2 diabetic or not diabetic patients affected by end-stage renal disease (ESRD). Material and Methods Thirty-six patients (20 affected by type 2 diabetes mellitus, and 16 not diabetic patients) were evaluated and underwent BHD dialysis, followed by HDF dialysis two days later. All patients underwent also glucose continuous monitoring system, using i Pro Continuous Glucose Monitor System (Medtronic MiniMed) starting just before the BHD, and ending five days later, two days after the HDF dialysis. Glycemic control was estimated as the mean blood glucose (MBG), the area under the glucose curve above 70mg/dl (AUC >70 ) or 180mg/dl (AUC >180 ), and the percentage of time above 70mg/dl (t >70 ) or 180mg/dl (t >180 ). Intraday glycemic variability was assessed as the standard deviation (SD), M value, and the mean amplitude of glycemic excursions (MAGE). Day-to-day glycemic variability was assessed as the mean of daily difference (MODD), that is the mean of the absolute difference among glucose values taken on 2 consecutive days at the same time. Results glycemic control was better with HDF: MBG, and AUC >180 were lower during HDF compared do BHD. We also observed a significant decrease of glycemic excursions during HDF dialysis: SD, M value, and the MAGE value were lower with HDF. The MODD value was significantly changed in BHD group, while no differences were recorded during HDF. Conclusion HDF seems to greater reduce glycemic excursions during the treatment compared to BHD.

Although mushroom poisoning is a rare cause of acute renal injury, in some cases it may lead to the development of a severe and irreversible renal failure. Orellanus syndrome is the most important example of organic renal damage related to mushroom consumption. It is caused by the ingestion of orellanine, the main toxin of different types of Cortinarius mushrooms (Cortinarius speciosissimus, C. orellanus, C. orellanoides, etc.), and it is characterized by progressive clinical phases with a predominant kidney involvement, finally requiring renal replacement therapy in about 10% of cases. Renal damage is often late and associated with a histological picture of interstitial nephritis. Diagnosis is essentially clinical and no specific therapy has been shown to be effective in preventing and treating renal damage. Here, we describe the case of a patient with mixed wild mushroom poisoning, presenting the typical clinical signs and course of the Orellanus syndrome. This case offers us the opportunity to review the main clinical features of this severe and little-known intoxication.

Toll-like receptor 4 (TLR4) promotes inflammation in hemodialysis patients (HD). A soluble form of extracellular TLR4 (sTLR4) has been recently characterized, which showed the ability to attenuate TLR4 signalling. In this study, we describe the sTLR4 profile in regular HD patients.In a cross-sectional study we enrolled forty prevalent HD patients (68.2 ± 16.3 years, twenty-five males) with a median dialysis vintage of 41 months. Nineteen patients were undergoing standard bicarbonate HD (BHD) and 21 patients on-line hemodiafiltration (HDF). Ten healthy sex-matched subjects constituted the controls (C).Before and after the HD session, serum was tested for sTLR4 levels by ELISA. Moreover, clinical and biochemical data were collected, including body mass index, albumin, and C-reactive protein (CRP) levels. Body composition was expressed as a 3-compartment model, providing lean tissue index and fat tissue index (FTI).Describe the profile of sTLR4 in HD patients, evaluating the correlations among sTLR4 levels and the main clinical characteristics, inflammatory and nutritional parameters.Patients with subclinical inflammation (i.e., high CRP levels without clinical symptomatology) presented higher sTLR4 levels (0.42 ± 0.25 ng/mL) with respect to both C and not inflamed HD patients (0.23 ± 0.19 ng/mL, P .05). There was a significant direct correlation between predialysis sTLR4 and body mass index, FTI (r = 0.55), and CRP levels (r = 0.52) and inverse correlation with lean tissue index and albumin (r = -0.4). In multivariate analysis, sTLR4 resulted directly associated with FTI (P = .038). Notably, sTLR4 levels resulted higher in bicarbonate hemodialysis versus hemodiafiltration (0.37 ± 0.18 vs. 0.19 ± 0.21 ng/mL, P .05).sTLR4 correlates with inflammatory and nutritional parameters, presenting as a new potential player in modulating subclinical inflammation in HD patients.

Kidney donation after circulatory death (DCD) is a less than ideal option to meet organ shortages. Hypothermic machine perfusion (HMP) with Belzer solution (BS) improves the viability of DCD kidneys, although the graft clinical course remains critical. Mesenchymal stromal cells (MSC) promote tissue repair by releasing extracellular vesicles (EV). We evaluated whether delivering MSC‐/MSC‐derived EV during HMP protects rat DCD kidneys from ischaemic injury and investigated the underlying pathogenic mechanisms. Warm ischaemic isolated kidneys were cold‐perfused (4 hrs) with BS, BS supplemented with MSC or EV. Renal damage was evaluated by histology and renal gene expression by microarray analysis, RT‐PCR. Malondialdehyde, lactate, LDH, glucose and pyruvate were measured in the effluent fluid. MSC‐/EV‐treated kidneys showed significantly less global ischaemic damage. In the MSC/EV groups, there was up‐regulation of three genes encoding enzymes known to improve cell energy metabolism and three genes encoding proteins involved in ion membrane transport. In the effluent fluid, lactate, LDH, MDA and glucose were significantly lower and pyruvate higher in MSC/EV kidneys as compared with BS, suggesting the larger use of energy substrates by MSC/EV kidneys. The addition of MSC/EV to BS during HMP protects the kidney from ischaemic injury by preserving the enzymatic machinery essential for cell viability and protects the kidney from reperfusion damage.

Several studies are increasingly underlying the biological role of vitamin E metabolites as bioactive compounds with anti-inflammatory, anti-proliferative and anti-atherogenic activity. A quantitative method for the simultaneous determination in human plasma and serum of vitamin E (α-tocopherol, α-T and γ-tocopherol, γ-T) and its cytochrome P-450 metabolites: 13'-hydroxychromanol (α-13'-OH), 13'-carboxychromanol (α-13'-COOH) and carboxyethyl hydroxychromanols (α-CEHC and γ-CEHC), was developed and validated. After enzymatic hydrolysis and deproteinization, the metabolites were extracted with a mixture of hexane/ methyl tertiary butyl ether (2/1, v/v). The separation was achieved by reversed phase chromatography and the analytes detected by a triple quadrupole mass analyser using electrospray ionization in positive mode (LC-MS/MS). α-T and γ-T were extracted separately without enzymatic hydrolysis. The analytes were quantified with the isotopic dilution method. After an extensive validation study (three levels in three different occasions for a total of 54 experiments), the procedure was successfully applied to the analysis of sera of healthy volunteers (before and after supplementation with α-T) and plasma of patients affected by chronic kidney disease. Finally, the structures of three unknown compounds found in blood and related to the long chain metabolites (α-13'-OH and α-13'-COOH) were further investigated using liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS).

Costimulatory pathways play a key role in immunity, providing the second signal required for a full activation of adaptive immune response. Different costimulatory families (CD28, TNF-related, adhesion and TIM molecules), characterized by structural and functional analogies, have been described. Costimulatory molecules modulate T cell activation, B cell function, Ig production, cytokine release and many other processes, including atherosclerosis. Patients suffering from renal diseases present significant alterations of the costimulatory pathways, which might make them particularly liable to infections. These alterations are further pronounced in patients undergoing kidney transplantation. In these patients, different costimulatory patterns have been related to distinct clinical features. The importance that costimulation has gained during the last years has led to development of several pharmacological approaches to modulate this critical step in the immune activation. Different drugs, mainly monoclonal antibodies targeting various costimulatory molecules (i.e. anti-CD80, CTLA-4 fusion proteins, anti-CD154, anti-CD40, etc.) were designed and tested in both experimental and clinical studies. The results of these studies highlighted some criticisms, but also some promising findings and now costimulatory blockade is considered a suitable strategy, with belatacept (a CTLA-4 fusion protein) being approved as the first costimulatory blocker for use in renal transplantation. In this review, we summarize the current knowledge on costimulatory pathways in the setting of kidney transplantation. We describe the principal costimulatory molecule families, their role and clinical significance in patients undergoing renal transplantation and the new therapeutic approaches that have been developed to modulate the costimulatory pathways.

This study was aimed to evaluate the effects on some inflammatory markers of two dialytic treatments [bicarbonate dialysis (BHD) and hemodiafiltration (HDF)] in patients with severe chronic kidney disease. We evaluated: blood glucose, homeostasis model assessment insulin resistance index, homocistein (Hcs), high sensitivity C-reactive protein (hs-CRP), fibrinogen, lipoprotein (a) [Lp(a)], metalloproteinases-2, and -9 (MMP-2 and MMP-9), and soluble receptor for advanced glycation end products (sRAGE). Considering the all sample, we observed a decrease of sRAGE with BHD, but not with HDF. Fibrinogen, MMP-2, and -9, Hs-CRP decreased after HDF, but not after BHD. In diabetics, blood glucose decreased after HDF dialysis, but not after BHD. Soluble receptor for advanced glycation end products obtained with HDF were higher compared to BHD. Fibrinogen, MMP-2, MMP-9, Hcs, Hs-CRP decreased, and ADN increased after HDF, these changes did not happen after BHD. Furthermore, sRAGE, and ADN were higher, and MMP-2 lower after HDF. In euglycemic patients, instead, MMP-2, MMP-9, and Hs-CRP decreased, and ADN increased with HDF, but not with BHD. We can conclude that hemodiafiltration seems to greater reduce inflammatory markers, and it could be more suitable for people with type 2 diabetes. Registration number: ClinicalTrials.gov NCT01049152.

Hepatocyte growth factor (HGF) is a pleiotropic cytokine which exerts a variety of effects on several cells, being involved in the regulation of many biological processes, such as inflammation, tissue repair, morphogenesis, angiogenesis, tumour propagation, immunomodulation of viral infections and cardio-metabolic activities. Patients undergoing regular hemodialysis (HD) present elevated levels of HGF, mainly due to the leukocyte activation associated with HD treatment. High HGF levels might account for specific clinical features of HD patients, i.e. mild liver damage in course of HCV-infection and high cardiovascular risk profile. Moreover, in patients with acute kidney injury, the induction of HGF may represent a crucial step to promote renal recovery, which can have important prognostic consequences in the short and long-term. In this review we discuss the mechanisms underlying HGF production in HD patients, the role of HGF in this particular patient population and the potential clinical implications derived from the study of HGF in HD patients.

Objective Advanced glycation end-products (AGEs) are proposed to influence inflammatory pathways and cardiovascular risk in chronic kidney disease (CKD). Dietary AGEs are believed to sustain circulating levels and toxicity in this condition. Design and Patients We investigated this aspect in a cross-sectional pilot study measuring levels of the AGE marker carboxymethyl lysine (CML) and fluorescent AGEs in the blood of pre-dialysis patients with CKD and hemodialysis (HD) patients (n = 10 each), and in a group of matched healthy controls (Ctr). Methods Plasma CML was measured by immuno-dot blot and fluorescent AGEs were determined by high-performace liquid chromatography (HPLC) analysis measuring the fluorescence of the cross-link pentosidine. The dietary intake of CML was assessed by dietary recall to trace total AGE intake in patients with CKD and the Ctr group. All the subjects included in the study were assessed for dietary intake while maintaining their usual diet. Main exclusion criteria for patients with CKD and HD were severe protein-caloric malnutrition and inflammation (measured by high sensitivity C-reactive protein and interleukin-6 levels). Results Plasma CML, as well as free and protein-bound fluorescent AGEs, significantly increased in CKD and even more in HD patients than that of the Ctr group. In patients with CKD, the average dietary intake of CML was less than half than that of the Ctr group (6 vs. 13 MU/day) and the lowered protein intake adopted spontaneously by these patients appear to explain this finding. Conclusions The results show that the intake of CML does not affect circulating levels of this as well as of other AGEs, in well nourished predialysis CKD patients.

Long-term hemodialysis treatment results in chronic monocyte activation with cytokine release. It generates Treg induction with potential immune dysfunction and associated clinical complications. Recent immunological data and preliminary clinical evidence suggest that synthetic polymers and vitamin E coated membranes are associated with a significant improvement in hemodialysis tolerance when compared to cellulose membranes. The aim of this review is to update cytokine release, T-cell polarization, and its clinical impact in patients under extracorporeal hemodialysis comparing traditional cellulose to synthetic/vitamin E coated membranes.

Autoimmunity to heat shock protein 60 (HSP60) has been related to atherosclerosis. Chlamydia pneumoniae (CP), the most studied infectious agent implicated in promoting atherosclerosis, produces a form of HSP60, which can induce an autoimmune response, due to high antigenic homology with human HSP60 (hHSP60). In this study, we evaluated the correlations among anti-hHSP60 antibodies, CP infection, and cardiovascular disease (CVD) in a high-risk population, such as patients undergoing hemodialysis (HD). Thirty-two patients (67.9 ± 13.9 years; male/female, 23:9) on regular HD were enrolled. Global absolute cardiovascular risk (GCR) was assessed using the Italian CUORE Project’s risk charts, which evaluate age, gender, smoking habits, diabetes, systolic blood pressure, and serum cholesterol. The occurrence of cardiovascular events during a 24-month follow-up was recorded. Seropositivity to CP and the presence of anti-hHSP60 antibodies were tested by specific enzyme-linked immunosorbent assays. Inflammation was assessed by measurement of C-reactive protein (CRP) serum levels. Fifteen healthy sex and age-matched (61.9 ± 9.5 years; male/female, 11:4) subjects were the control group. Fifteen of 32 patients resulted seropositive for CP. CP + patients were older than CP−, while they did not differ for GCR, CRP, and dialytic parameters. CVD incidence was significantly higher in CP+ (9 CP+ vs 2 CP−, p

Epidemiology data predict that by the year 2025, diabetes will affect about 380 million people worldwide with a significant increase in patients with chronic renal disease progressing to hemodialysis. Diabetes-related peripheral vascular disease is a major risk factor for vascular access failure in patients on extracorporeal hemodialysis. Although peritoneal dialysis is a valid option for diabetics, peritonitis is still a main complication for these patients. We report the case of a 71-year-old type 2 diabetes patient treated by subcutaneous insulin, undergoing automated peritoneal dialysis (APD) who developed peritonitis and bloodstream infection by Ochrobactrum anthropi (O. anthropi). The patient was initially shifted to continuous ambulatory peritoneal dialysis (CAPD) and treated with intraperitoneal cefotaxime and gentamicin. According to antibiogram, cefotaxime was discontinued but lasting gentamicin. Within 48 h from admission, clear peritoneal effluent was observed with reduction in white blood cells count from 580/mm³ 77.9% neutrophils to less than 10/mm³. Prompt regression of infection without catheter removal and no relapse after over 7-month follow-up allowed supposing that O. anthropi did not colonized peritoneal catheter. O. anthropi is an emerging cause of nosocomial infection in immunocompromised patients. Cases of such infection in patients undergoing CAPD and hemodialysis have been already described. However, this is the first reported case of O. anthropi in a patient undergoing APD.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Mycophenolic acid (MPA) is a potent, selective and reversible inhibitor of inosine 5′-monophosphate dehydrogenase (IMPDH), the rate-limiting enzyme for de novo guanosine triphosphate biosynthesis. • The large IMPDH interindividual variability could be responsible for the differences in therapeutic effects and side-effects observed with MPA. • Induction of IMPDH activity has been observed in whole blood during immunosuppressive therapy. WHAT THIS STUDY ADDS • Our data were acquired in long-term mycophenolate mofetil-treated renal transplant recipients on different combinations of immunosuppressive agents (ciclosporin, tacrolimus, sirolimus) and with different treatment duration (up to 8.8 years post transplant). • The increasing trend in IMPDH activity that we observed throughout our 12-month observation period was significantly higher in rejecting than in nonrejecting subjects. AIMS Long-term mycophenolate mofetil (MMF) therapy may induce inosine 5′-monophosphate dehydrogenase (IMPDH) activity in peripheral blood mononuclear cells (PBMCs), thus decreasing MMF immunosuppressive properties. Pharmacodynamic monitoring was used to investigate whether biological activity is altered after long-term therapy. METHODS IMPDH activity was measured in PBMC samples from 54 stable kidney transplant patients, already on MMF (for at least 3 months), before (t0) and 2 h after (t2) MMF morning dose administration; levels were monitored for up to 15 months, together with total mycophenolic acid (MPA) and free MPA concentrations. RESULTS During the 15 months' monitoring, t0 IMPDH activity in transplant recipients increased from 5.9 ± 3.7 nmol h−1 mg−1[95% confidence interval (CI) 4.9, 6.9] to 9.0 ± 3.9 nmol h−1 mg−1 (95% CI 7.2, 10.8), with an intra- and interpatient variability of 28% and 42%. Five patients experienced acute rejection during the follow-up: t0 IMPDH activity was increased during rejection vs. nonrejection, and the trend was significantly higher in rejecting than in nonrejecting subjects for the whole monitoring period. CONCLUSIONS Even though a correlation has been found between IMPDH activity and rejection, its efficacy as a predictive tool in long-term transplant outcomes may be affected by high interpatient variability; on the other hand, continuous monitoring of the IMPDH trend could make an effective prognostic parameter of rejection. Other trials also including pre-transplant data on both IMPDH expression and activity are warranted to better assess their role as biomarkers for MPA effect in clinical practice.

While sirolimus (SRL) is thought to be a non-nephrotoxic agent, cyclosporine A (CsA) toxicity is a serious problem in kidney transplantation. We compared the effects of the two drugs on T-helper (Th) subsets in kidney transplant patients. We examined 24 first cadaver kidney recipients equally randomized to receive SRL/mycophenolate mofetil (MMF)/methylprednisolone (MP), or cyclosporine with either MMF or MP. The Th1 and Th2 subsets in peripheral blood were separated based on their production of interferon-gamma (INFgamma) or interleukin (IL)-4/IL-5. The lymphocytes were stimulated with phytohemoagglutinin or with allogenic CD3-depeted and irradiated antigen-presenting cells. Furthermore, the conversion potential of Th0 to Th1 was determined by measuring IL-12 and IL-18 levels after lipopolysaccharide challenge. When peripheral blood lymphocytes taken from SRL-treated patients were stimulated by phytohemoagglutinin, there were significantly lower INFgamma-producing cells compared with the lymphocytes taken from patients treated with CsA. The number of IL-4/IL-5-producing cells did not differ among the patient groups. Release of IL-12 but not IL-18 from peripheral lymphocytes following treatment with lipopolysaccharide was significantly lower in the SRL-treated patients. These results show that compared with CsA, SRL caused a significant decrease in the Th1 lymphocyte subset associated with a significant reduction of IL-12 release.

Macrophage-stimulating protein (MSP) is a scatter factor that causes cell proliferation and migration, and receptor origin nantaise (RON) is its receptor. RON is expressed in macrophages and mesangial cells, and MSP is produced by renal tubular cells. This study investigated whether MSP/RON participate in the pathogenesis of anti-Thy 1 nephritis, a glomerular disease that is characterized by invasion of circulating monocytes into glomeruli and migration and proliferation of mesangial cells. In vivo, renal function and histopathology were studied in rats that had anti-Thy 1 disease and were untreated and treated with a neutralizing anti-MSP antibody. In vitro, whether monocytes express RON and whether MSP has a chemotactic effect on monocytes were studied. In vivo, in anti-Thy 1 disease, MSP was expressed de novo in glomeruli, and neutralization of MSP attenuated the rise in serum creatinine and proteinuria, stopped glomerular neutrophil and monocyte influx, protected from glomerular injury, and lessened mesangial cell overgrowth. In vitro, unstimulated monocytes did not express RON, but the stimulation with LPS induced de novo RON expression. LPS-stimulated monocytes were attracted by MSP. These results demonstrate a pathogenic role of the MSP/RON system in anti-Thy 1 nephritis.

Progressive hemodialysis is based on the simple idea of adjusting its dose according to residual renal function (RRF). The progressive, infrequent paradigm is slowly gaining a foothold among nephrologists, despite a lot of skepticism in the scientific world. Given the importance of RRF preservation in conservative therapy, it seems a contradiction to ignore the contribution of RRF when patients initiate hemodialysis (HD), especially when it is routinely considered with peritoneal dialysis. While a three-times-weekly HD regimen is broadly considered the standard starting regimen for new patients, twice-weekly HD has been used in selected patients and is currently a common practice in South-East Asia. Small studies indicate that a once-weekly HD regimen may be a viable starting option as well. Progressive hemodialysis still requires validation, yet it is promising. We share the belief that a randomized clinical trial to investigate progressive hemodialysis is much needed, but we also strongly recommend including a once-weekly HD starting dose as part of any such investigation.

Regulatory T cells (Tregs), defined as CD4+CD25+highFoxP3+CD127- cells, could promote tolerance in renal transplantation (Tx). In an open-label, randomized, controlled trial 62 de-novo Tx recipients received induction with basiliximab and cyclosporine A (CsA) for the first month after Tx and then were assigned to treatment with sirolimus (SRL) or CsA and followed up for 2 years. The primary endpoint was to evaluate the effects of induction and maintenance treatments on circulating Tregs, while the secondary endpoint was the assessment of Treg renal infiltration and the relationship between Treg count and clinical outcomes. There were no significant differences in either circulating or tissue Treg number between the two groups. At 1 month post-Tx, all patients presented a profound Treg depletion, followed by a significant increase in Tregs that resulted stable during the follow-up. The same trend was also observed for non-activated Tregs (CD69-) and for other immunocompetent cells (CD4+ and CD8+ T cells, B cells and NK cells). Moreover, the Treg count did not correlate either with renal function or with acute rejection and graft loss. Initial immunosuppression is crucial to regulate circulating Tregs, regardless of subsequent immunosuppressive maintenance regimens. Strategies aiming to promote tolerance should consider the effects of different induction regimens.

Since the early description of Randall plaques in 1937, studies of the pathogenesis of stone formation mainly focused on the chemistry involving salt precipitation and crystallization, rather than tubular and interstitial medullar mechanisms of calcium concentration and supersaturation. In 2003, Bushinsky published a suggestive and inspiring sequence of events aimed to show that the basement membrane of the thin limb of the loop of Henle can be the first site of nucleation, as recently shown by the impressive work by Evan et al. The aim of this minireview is to verify the consistency of the Evan and Bushinsky theory with the current literature in the field.

Heparin has remained the most commonly used anticoagulant in hemodialysis patients (HD). Its use is usually safe but, in some cases, important adverse effects can occur. Heparin-induced thrombocytopenia (HIT) is an immuno-mediated condition due to the formation of PF4/heparin/IgG complex leading to the activation of platelets and coagulative cascade. The consequent prothrombotic hypercoagulable state may cause venous or arterial thrombosis, skin gangrene and acute platelet activation syndrome. Clinical and laboratory findings may be suggestive for HIT, but formal diagnosis requires the demonstration of the presence of circulating antibodies. Clinical management is complex including the withdrawal of any form of heparin and the administration of anticoagulants. In addition, since anticoagulation is routinely required to prevent clotting of the dialysis lines and membranes, in HD patients presenting HIT it is mandatory to establish heparin-free anticoagulation strategies. Thus, the use of citrate, direct thrombin inhibitors or eparinods have been proposed as alternative anticoagulation approaches in HIT. Here, we review the most important pathogenic factors and clinical features of HIT occurring in HD patients.

T-helper (Th) lymphocytes consist of Th1 and Th2 subsets. Th1 cells are effectors of cell-mediated immunity and secrete interferon-gamma (IFN-gamma), which recruits new Th1 cells in cooperation with interleukin-12 (IL-12; produced by monocytes) and inhibits Th2 differentiation. Th2 cells produce IL-4 and IL-10, which inhibit IFN-gamma secretion and cell immunity. We investigated whether the impaired immune response in uremia is associated with an altered balance of Th1/Th2. Peripheral-blood mononuclear cells (PBMCs) were collected from patients with chronic renal failure (CRF) on conservative treatment (CRF patients), patients with end-stage renal disease (ESRD) on regular hemodialysis therapy (ESRD-HD patients), and healthy controls (CON). CD4(+) cells were isolated from PBMCs by negative selection using a magnetic labeling system. PBMCs and purified CD4(+) cells were cultured in Iscove's medium and Iscove's medium plus mitogens (phytohemagglutinin and lipopolysaccharide). IFN-gamma, IL-12, IL-4, and IL-10 were measured in supernatant. The constitutive release of IL-4 and IL-10 by PBMCs and CD4(+) cells of CRF and ESRD-HD patients was increased by five to eight times in comparison with CON (P < 0.001). Constitutive IFN-gamma release by PBMCs of ESRD-HD patients was undetectable, although they secreted an increased amount of IL-12. Mitogen-stimulated release of IFN-gamma by PBMCs and CD4(+) cells of CRF and ESRD-HD patients was blunted (average PBMCs: CON, 115.8 pg/2 x10(6) cells; CRF, 81.8 pg/2 x10(6) cells; ESRD-HD, 9.3 pg/2 x10(6) cells; CD4(+) cells: CON, 358.0 pg/5 x 10(5) cells; CRF, 165.4 pg/5 x 10(5) cells; ESRD-HD, 43.5 pg/5 x 10(5) cells). The ability of PBMCs of ESRD-HD patients to secrete IFN-gamma was recovered after IL-4 and IL-10 neutralization. Uremia is associated with a prevalence of Th1 over Th2 cells and a configuration of cytokine network that depresses cell-mediated immunity.

Sibutramine, a serotonin reuptake inhibitor, currently is used in treatment of obesity. The known side effects of sibutramine, ie, hypertension and tachycardia, depend on its adrenergic and serotoninergic effects. We describe a case of life-threatening hyponatremia associated with sibutramine use in an obese woman. We hypothesize that sibutramine, through its effect on neurotransmitters, may induce antidiuretic hormone secretion and lead to a syndrome of inappropriate antidiuretic hormone secretion. We advise careful monitoring of water-electrolytic balance during sibutramine therapy.

Hepatocyte growth factor (HGF) is a potent mitogen for tubular cells. Experimental injury to the kidney is associated with HGF release both locally and by distant organs stimulated by circulating ‘injurins’. In this study, the serum HGF concentration was measured in patients with acute renal failure (ARF). Normal subjects and chronic renal failure patients either not on dialysis or on regular dialysis treatment served as controls. Human mesangial cells were incubated with sera from ARF patients and controls. The serum HGF concentration was strikingly increased in ARF patients (478 ± 68 ng/dl) and was normal in chronic renal failure patients not on dialysis (20 ± 3 ng/dl) and in those on regular dialysis treatment (25 ± 3 ng/dl). Serum of ARF patients strongly stimulated HGF release from mesangial cells (1,384 ± 55 ng/ml) in comparison with normal serum (67 ± 10 ng/ml). These results indicate that in ARF HGF participates in tubular repair both as an endocrine factor, released in the circulation, and as a paracrine substance, diffusing to the tubules from the mesangium.