Gut microbiota and fecal volatilome profile inspection in metabolically healthy and unhealthy obesity phenotypes.

BACKGROUND: People with metabolically healthy (MHO) and metabolically unhealthy obesity (MUO) differ for the presence or absence of cardio-metabolic complications, respectively. OBJECTIVE: Based on these differences, we are interested in deepening whether these obesity phenotypes could be linked to changes in microbiota and metabolome profiles. In this respect, the overt role of microbiota taxa composition and relative metabolic profiles is not completely understood. At this aim, biochemical and nutritional parameters, fecal microbiota, metabolome and SCFA compositions were inspected in patients with MHO and MUO under a restrictive diet regimen with a daily intake ranging from 800 to 1200 kcal. METHODS: Blood, fecal samples and food questionnaires were collected from healthy controls (HC), and an obese cohort composed of both MHO and MUO patients. Most impacting biochemical/anthropometric variables from an a priori sample stratification were detected by applying a robust statistics approach useful in lowering the background noise. Bacterial taxa and volatile metabolites were assessed by qPCR and gas chromatography coupled with mass spectrometry, respectively. A targeted GC-MS analyses on SCFAs was also performed. RESULTS: Instructed to follow a controlled and restricted daily calorie intake, MHO and MUO patients showed differences in metabolic, gut microbial and volatilome signatures. Our data revealed higher quantities of specific pro-inflammatory taxa (i.e., Desulfovibrio and Prevotella genera) and lower quantities of Clostridium coccoides group in MUO subset. Higher abundances in alkane, ketone, aldehyde, and indole VOC classes together with a lower amount of butanoic acid marked the faecal MUO metabolome. CONCLUSIONS: Compared to MHO, MUO subset symptom picture is featured by specific differences in gut pro-inflammatory taxa and metabolites that could have a role in the progression to metabolically unhealthy status and developing of obesity-related cardiometabolic diseases. The approach is suitable to better explain the crosstalk existing among dysmetabolism-related inflammation, nutrient intake, lifestyle, and gut dysbiosis.

Ozone eliminates novel coronavirus Sars-CoV-2 in mucosal samples.

Recent investigations have shown that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is able to resist on the surfaces and that the diffusion occurs through droplets that can remain suspended in the air as an aerosol. The ozone generated in situ from oxygen is an active ingredient with a 'biocidal' action, but little is known about its capacity to inactivate specifically SARS-CoV-2. Here we show, for the first time, the efficiency of the ozone treatment to neutralize the SARS-CoV-2 present in nasopharynx secretion samples with high viral load. Our data show that ozone is effective in SARS-CoV-2 elimination.

Severe acute respiratory syndrome coronavirus 2 may exploit human transcription factors involved in retinoic acid and interferon-mediated response: a hypothesis supported by an in silico analysis.

The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing coronavirus disease 2019 (COVID-19), resulting in acute respiratory disease, is a worldwide emergency. Because recently it has been found that SARS-CoV is dependent on host transcription factors (TF) to express the viral genes, efforts are required to understand the molecular interplay between virus and host response. By bioinformatic analysis, we investigated human TF that can bind the SARS-CoV-2 sequence and can be involved in viral transcription. In particular, we analysed the key role of TF involved in interferon (IFN) response. We found that several TF could be induced by the IFN antiviral response, specifically some induced by IFN-stimulated gene factor 3 (ISGF3) and by unphosphorylated ISGF3, which were found to promote the transcription of several viral open reading frame. Moreover, we found 22 TF binding sites present only in the sequence of virus infecting humans but not bat coronavirus RaTG13. The 22 TF are involved in IFN, retinoic acid signalling and regulation of transcription by RNA polymerase II, thus facilitating its own replication cycle. This mechanism, by competition, may steal the human TF involved in these processes, explaining SARS-CoV-2's disruption of IFN-I signalling in host cells and the mechanism of the SARS retinoic acid depletion syndrome leading to the cytokine storm. We identified three TF binding sites present exclusively in the Brazilian SARS-CoV-2 P.1 variant that may explain the higher severity of the respiratory syndrome. These data shed light on SARS-CoV-2 dependence from the host transcription machinery associated with IFN response and strengthen our knowledge of the virus's transcription and replicative activity, thus paving the way for new targets for drug design and therapeutic approaches.

Effective and safe off-label use of caplacizumab treatment in a middle-aged obese male.

This study shows clinical efficacy and safety profile of an off-label use of caplacizumab for the treatment of immune-mediated thrombotic thrombocytopenic purpura in a middle-aged obese male patient manifesting aphasia, weakness and unconsciousness. Routine blood tests revealed haemolytic anaemia, severe thrombocytopenia (platelet count=20×109/L) and moderate creatinine increase. Diagnosis was based on the clinical judgement and laboratory determinations (undetectable ADAMTS13 activity and presence of anti-ADAMTS13 antibodies). The patient underwent plasma-exchange and an adjunctive treatment with prednisone (1mg/Kg/day), but the occurrence of a refractory and exacerbated form of disease suggested also using rituximab (375mg/m2 weekly for 4 weeks) and caplacizumab as salvage treatments. The caplacizumab was given at 10mg/day subcutaneously without the first intravenous bolus. Because von Willebrand factor inhibition, platelet count recovery and remission of symptoms were achieved, use of caplacizumab with this scheme appeared to be as effective as the approved one. Although this is an off-label use, this case highlights the potential of this new treatment, in terms of drug's efficacy and safety.

Hyperuricemia and Risk of Cardiovascular Outcomes: The Experience of the URRAH (Uric Acid Right for Heart Health) Project.

The latest European Guidelines of Arterial Hypertension have officially introduced uric acid evaluation among the cardiovascular risk factors that should be evaluated in order to stratify patient's risk. In fact, it has been extensively evaluated and demonstrated to be an independent predictor not only of all-cause and cardiovascular mortality, but also of myocardial infraction, stroke and heart failure. Despite the large number of studies on this topic, an important open question that still need to be answered is the identification of a cardiovascular uric acid cut-off value. The actual hyperuricemia cut-off (> 6 mg/dL in women and 7 mg/dL in men) is principally based on the saturation point of uric acid but previous evidence suggests that the negative impact of cardiovascular system could occur also at lower levels. In this context, the Working Group on uric acid and CV risk of the Italian Society of Hypertension has designed the Uric acid Right for heArt Health project. The primary objective of this project is to define the level of uricemia above which the independent risk of CV disease may increase in a significantly manner. In this review we will summarize the first results obtained and describe the further planned analysis.

Quasi-3D morphology and modulation of focal adhesions of human adult stem cells through combinatorial concave elastomeric surfaces with varied stiffness.

In vivo cell niches are complex architectures that provide a wide range of biochemical and mechanical stimuli to control cell behavior and fate. With the aim to provide in vitro microenvironments mimicking physiological niches, microstructured substrates have been exploited to support cell adhesion and to control cell shape as well as three dimensional morphology. At variance with previous methods, we propose a simple and rapid protein subtractive soft lithographic method to obtain microstructured polydimethylsiloxane substrates for studying stem cell adhesion and growth. The shape of adult renal stem cells and nuclei is found to depend predominantly on micropatterning of elastomeric surfaces and only weakly on the substrate mechanical properties. Differently, focal adhesions in their shape and density but not in their alignment mainly depend on the elastomer stiffness almost regardless of microscale topography. Local surface topography with concave microgeometry enhancing adhesion drives stem cells in a quasi-three dimensional configuration where stiffness might significantly steer mechanosensing as highlighted by focal adhesion properties.

Impact of 3 Major Maintenance Immunosuppressive Protocols on Long-term Clinical Outcomes: Result of a Large Multicenter Italian Cohort Study Including 5635 Renal Transplant Recipients.

BACKGROUND: Although optimization of immunosuppressive schemes in renal transplantation have minimized acute posttransplant complications, long-term outcomes are still not optimal and most of the chronic graft damage is drug-related. Therefore, to define the best long-term maintenance immunosuppressive regimen is of major importance in renal transplantation. To assess this objective, we undertook a large, multicenter cohort study in Italy. METHODS: We retrospectively analyzed data of 5635 patients (enrolled from 1983 to 2012) and we assessed the impact of 3 major immunosuppressive regimens (calcineurin inhibitors+antimetabolites+corticosteroids [CNI+ANT+CS] vs CNI+mammalian target-of-rapamycin (mTOR) inhibitors+CS [CNI+mTOR-I+CS] vs CNI+CS) on long-term clinical outcomes by employing several statistical algorithms. RESULTS: The overall difference in the incidence of outcome over time was not statistically different within the first 5 years of follow-up (P = .13); however, it became significant at 10 years and 20 years (P < .01), with the CNI+CS group showing the lowest cumulative incidence of outcome. Compared with the CNI+ANT+CS group, the CNI+mTOR-I+CS group patients had a significantly higher risk of outcome (hazard ratio [HR], 1.30; P = .024); the difference remained significant and even increased in magnitude after adjustment for potential confounders (HR, 1.38; P = .006). Similarly, patients in the CNI+CS group had a significantly higher risk of the outcome (HR, 1.64; P < .001). CONCLUSION: Our data confirm that CNI+ANT+CS is the "gold standard" therapy in renal transplantation, but, whenever required, the introduction of mTOR-Is instead of ANT may not dramatically modify major clinical outcomes. The use of mTOR-I could be a valuable pharmacologic tool to minimize CNI complications and insure adequate immunosuppression.

Intra-dialytic blood oxygen saturation (SO2): association with dialysis hypotension (the SOGLIA Study).

BACKGROUND: Intradialytic hypotension (IDH) has a dramatic impact on the main outcomes of dialysis patients. Early warning of hemodynamic worsening during dialysis would enable preventive measures to be taken. Blood oxygen saturation (SO2) is used for hemodynamic monitoring in the critical care setting and may provide useful information about IDH onset. AIM: To evaluate whether short- and medium-term variations in the SO2 signal (ST-SO2var, MT-SO2var,) during dialysis are a predictor of IDH. METHODS: In this 3-month observational cohort study, 51 hypotension-prone chronic hemodialysis (HD) patients, with vascular access by arteriovenous fistula (AVF) or central venous catheter (CVC), were enrolled. Continuous non-invasive blood SO2 was monitored (fc = 0.2 Hz) by an optical sensor on the arterial line of the extracorporeal circulation; blood pressure (every 30 min), symptoms and their time of appearance were noted. Predictive power of IDH was expressed by the area under curve (AUC) sensitivity and specificity based on intradialytic variations in SO2. RESULTS: A total of 1290 HD sessions were analyzed. Overall, off-line ST-SO2var analysis proved able to correctly predict IDH in 67 % of the sessions where IDH occurred. The best predictive performance was found in the presence of highly arterialized AVF (SO2 > 95 %) (75 % sensitivity; AUC 0.825; p < 0.05). On the contrary, in sessions with CVC, IDH prediction proved more efficient by MT-SO2var (AUC 0.575; p = 0.01). CONCLUSIONS: Intradialytic SO2 variability could be a valid parameter to detect in advance the hemodynamic worsening that precedes IDH. Appropriate timely intervention could help prevent IDH onset.

Complement Modulation of Anti-Aging Factor Klotho in Ischemia/Reperfusion Injury and Delayed Graft Function.

Klotho is an anti-aging factor mainly produced by renal tubular epithelial cells (TEC) with pleiotropic functions. Klotho is down-regulated in acute kidney injury in native kidney; however, the modulation of Klotho in kidney transplantation has not been investigated. In a swine model of ischemia/reperfusion injury (IRI), we observed a remarkable reduction of renal Klotho by 24 h from IRI. Complement inhibition by C1-inhibitor preserved Klotho expression in vivo by abrogating nuclear factor kappa B (NF-kB) signaling. In accordance, complement anaphylotoxin C5a led to a significant down-regulation of Klotho in TEC in vitro that was NF-kB mediated. Analysis of Klotho in kidneys from cadaveric donors demonstrated a significant expression of Klotho in pre-implantation biopsies; however, patients affected by delayed graft function (DGF) showed a profound down-regulation of Klotho compared with patients with early graft function. Quantification of serum Klotho after 2 years from transplantation demonstrated significant lower levels in DGF patients. Our data demonstrated that complement might be pivotal in the down-regulation of Klotho in IRI leading to a permanent deficiency after years from transplantation. Considering the anti-senescence and anti-fibrotic effects of Klotho at renal levels, we hypothesize that this acquired deficiency of Klotho might contribute to DGF-associated chronic allograft dysfunction.

Acquired lecithin:cholesterol acyltransferase deficiency as a major factor in lowering plasma HDL levels in chronic kidney disease.

OBJECTIVES: It has been suggested that a low plasma high-density lipoprotein cholesterol (HDL-C) level contributes to the high cardiovascular disease risk of patients with chronic kidney disease (CKD), especially those undergoing haemodialysis (HD). The present study was conducted to gain further understanding of the mechanism(s) responsible for the low HDL-C levels in patients with CKD and to separate the impact of HD from that of the underlying CKD. METHODS: Plasma lipids and lipoproteins, HDL subclasses and various cholesterol esterification parameters were measured in a total of 248 patients with CKD, 198 of whom were undergoing HD treatment and 40 healthy subjects. RESULTS: Chronic kidney disease was found to be associated with highly significant reductions in plasma HDL-C, unesterified cholesterol, apolipoprotein (apo)A-I, apoA-II and LpA-I:A-II levels in both CKD cohorts (with and without HD treatment). The cholesterol esterification process was markedly impaired, as indicated by reductions in plasma lecithin:cholesterol acyltransferase (LCAT) concentration and activity and cholesterol esterification rate, and by an increase in the plasma preß-HDL content. HD treatment was associated with a further lowering of HDL levels and impaired plasma cholesterol esterification. The plasma HDL-C level was highly significantly correlated with LCAT concentration (R = 0.438, P < 0.001), LCAT activity (R = 0.243, P < 0.001) and cholesterol esterification rate (R = 0.149, P = 0.031). Highly significant correlations were also found between plasma LCAT concentration and levels of apoA-I (R = 0.432, P < 0.001), apoA-II (R = 0.275, P < 0.001), LpA-I (R = 0.326, P < 0.001) and LpA-I:A-II (R = 0.346, P < 0.001). CONCLUSION: Acquired LCAT deficiency is a major cause of low plasma HDL levels in patients with CKD, thus LCAT is an attractive target for therapeutic intervention to reverse dyslipidaemia, and possibly lower the cardiovascular disease risk in these patients.

Sclerostin and Dickkopf-1 in post-menopausal renal allograft recipients.

BACKGROUND: Age, pre-existing renal osteodistrophy, impaired renal function, and chronic use of immunosuppressive drugs are the main factors involved in the onset and development of bone metabolism disturbances and skeletal alterations occurring after renal transplantation. However, at the state of the art, no reports have analyzed the additional post-menopausal physiological mechanisms associated with the onset and development of bone complications in renal transplant recipients. METHODS: We measured by means of molecular strategies (enzyme-linked immunoassay, chemiluminescence) the serum levels of Sclerostin and Dickkopf-1 (DKK1), two major antagonists of the Wnt/ß-catenin pathway, and several bone resorption/formation biomarkers (N-terminal procollagen type 1, bone-specific alkaline phosphatase, and serum C-terminal telopeptides of type I collagen) in 19 post-menopausal kidney transplant patients and 12 post-menopausal chronic kidney disease patients (CKD group) matched for age and renal function. RESULTS: Our results showed that the levels of both Wnt antagonists were similar in the two study groups (P=.15 and .96, respectively). Additionally, no correlation was found between Sclerostin and DKK1 serum levels in all patients included in the study (R2=0.03, P=.2). After statistical analysis, we found no differences in the bone resorption/formation biomarkers between renal transplant and CKD patients. Multivariate analysis showed that Sclerostin levels were significantly positively correlated with serum phosphorus levels (P=.008) and inversely correlated with renal function (P=.026). Surprisingly, no significant correlation was found between all the analyzed demographic and clinical parameters and DKK1. CONCLUSIONS: Our study demonstrated for the first time that renal transplantation per se and immunosuppressive treatments do not represent additional factors contributing to bone metabolic/biochemical alterations in post-menopausal women. However, our results emphasized that a better preservation of the graft function could significantly slow down the progression of bone metabolic deregulations and prevent clinical bone complications.

Complement-dependent NADPH oxidase enzyme activation in renal ischemia/reperfusion injury.

NADPH oxidase plays a central role in mediating oxidative stress during heart, liver, and lung ischemia/reperfusion injury, but limited information is available about NADPH oxidase in renal ischemia/reperfusion injury. Our aim was to investigate the activation of NADPH oxidase in a swine model of renal ischemia/reperfusion damage. We induced renal ischemia/reperfusion in 10 pigs, treating 5 of them with human recombinant C1 inhibitor, and we collected kidney biopsies before ischemia and 15, 30, and 60 min after reperfusion. Ischemia/reperfusion induced a significant increase in NADPH oxidase 4 (NOX-4) expression at the tubular level, an upregulation of NOX-2 expression in infiltrating monocytes and myeloid dendritic cells, and 8-oxo-7,8-dihydro-2'-deoxyguanosine synthesis along with a marked upregulation of NADPH-dependent superoxide generation. This burden of oxidative stress was associated with an increase in tubular and interstitial expression of the myofibroblast marker α-smooth muscle actin (α-SMA). Interestingly, NOX-4 and NOX-2 expression and the overall NADPH oxidase activity as well as α-SMA expression and 8-oxo-7,8-dihydro-2'-deoxyguanosine synthesis were strongly reduced in C1-inhibitor-treated animals. In vitro, when we incubated tubular cells with the anaphylotoxin C3a, we observed an enhanced NADPH oxidase activity and α-SMA protein expression, which were both abolished by NOX-4 silencing. In conclusion, our findings suggest that NADPH oxidase is activated during ischemia/reperfusion in a complement-dependent manner and may play a potential role in the pathogenesis of progressive renal damage in this setting.

Effects of ischemia-reperfusion injury in kidney transplantation: risk factors and early and long-term outcomes in a single center.

INTRODUCTION: Ischemia-reperfusion injury (IRI) causes a high rate of delayed graft function (DGF), the most frequent complication in the immediate postoperative period after cadaveric donor kidney transplantation. Herein we evaluated the impact of donor and recipient characteristics on DGF development in terms of the incidence of acute rejection episodes, hospital stay, renal function, and long-term graft and patient survivals. MATERIALS AND METHODS: Between February 1998 and July 2011, 761 patients underwent cadaveric donor kidney transplantations. DGF was defined as the need for dialysis in the first week. Patients were subdivided according to initial graft function as immediate graft function (IGF) or DGF. RESULTS: DGF observed in 241 patients (31.6%) was associated independently with expanded criteria donors, extended cold ischemia time, Karpinsky histological score, and prior dialysis duration both univariate and multivariate analysis. The incidence of acute rejection episodes was 18.1% among the DGF group versus 1.3% in the IGF group (P < .01). DGF significantly reduced both graft and patient survivals at 6, 12, 36, and 60 months. CONCLUSION: DGF was responsible for a longer hospital stay, worse early and long-term renal function, a higher incidence of acute rejection episodes as well as reduced graft and patient survivals.

Advanced age is not an exclusion criterion for kidney transplantation.

INTRODUCTION: Renal transplantation in patients older than 60 years has long been regarded with skepticism owing to the increased risk of complications although, as compared with dialysis treatment, a graft seems to improve not only the quality of life but also long-term patient survival. This study sought to analyze the impact of recipient age older than 60 years on patient and graft outcomes. MATERIALS AND METHODS: We retrospectively investigated the outcomes of 761 kidney transplant recipients from cadaveric donors performed between February 1998 and July 2011. While 69 subjects were at least 60 years of age (group A), 692 were younger than 60 years (group B) at the time of transplantation. RESULT: Mean follow-up was 60.1 ± 38.5 months. Delayed graft function (DGF) requiring dialysis was observed in 36 group A (52.1%) and 205 group B (29.6%) subjects (P = .001). However, there were also significant differences between group A and group B in terms of mean donor age (60.3 vs 44.6 years; P < .001) and mean donor estimated creatinine clearance (57.8 vs 83.4 mL/min; P < .001). There were no significant differences in death-censored graft survival between the two groups, but elderly patients experienced worse survival (P = .0005). The most common causes of patient death were myocardial infarction, other cardiovascular complications, and tumors. CONCLUSION: Kidney transplantation is a good option for elderly recipients with end-stage renal disease, providing long graft survival and a good quality of life, although these patients are more likely to develop cancer or cardiovascular disease. Our findings suggested that older patients should not be excluded a priori from transplantation, but meticulous screening for cancer and heart disease should be always be performed to improve outcomes.