Publications by authors named "Elisa Benetti"

84 Publications

Shorter androgen receptor polyQ alleles protect against life-threatening COVID-19 disease in European males.

EBioMedicine 2021 Feb 26;65:103246. Epub 2021 Feb 26.

Medical Genetics, University of Siena, Italy; Med Biotech Hub and Competence Center, Department of Medical Biotechnologies, University of Siena, Italy; Genetica Medica, Azienda Ospedaliero-Universitaria Senese, Italy.

Background: While SARS-CoV-2 similarly infects men and women, COVID-19 outcome is less favorable in men. Variability in COVID-19 severity may be explained by differences in the host genome.

Methods: We compared poly-amino acids variability from WES data in severely affected COVID-19 patients versus SARS-CoV-2 PCR-positive oligo-asymptomatic subjects.

Findings: Shorter polyQ alleles (≤22) in the androgen receptor (AR) conferred protection against severe outcome in COVID-19 in the first tested cohort (both males and females) of 638 Italian subjects. The association between long polyQ alleles (≥23) and severe clinical outcome (p = 0.024) was also validated in an independent cohort of Spanish men <60 years of age (p = 0.014). Testosterone was higher in subjects with AR long-polyQ, possibly indicating receptor resistance (p = 0.042 Mann-Whitney U test). Inappropriately low serum testosterone level among carriers of the long-polyQ alleles (p = 0.0004 Mann-Whitney U test) predicted the need for intensive care in COVID-19 infected men. In agreement with the known anti-inflammatory action of testosterone, patients with long-polyQ and age ≥60 years had increased levels of CRP (p = 0.018, not accounting for multiple testing).

Interpretation: We identify the first genetic polymorphism that appears to predispose some men to develop more severe disease. Failure of the endocrine feedback to overcome AR signaling defects by increasing testosterone levels during the infection leads to the polyQ tract becoming dominant to serum testosterone levels for the clinical outcome. These results may contribute to designing reliable clinical and public health measures and provide a rationale to test testosterone as adjuvant therapy in men with COVID-19 expressing long AR polyQ repeats.

Funding: MIUR project "Dipartimenti di Eccellenza 2018-2020" to Department of Medical Biotechnologies University of Siena, Italy (Italian D.L. n.18 March 17, 2020) and "Bando Ricerca COVID-19 Toscana" project to Azienda Ospedaliero-Universitaria Senese. Private donors for COVID-19 research and charity funds from Intesa San Paolo.
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http://dx.doi.org/10.1016/j.ebiom.2021.103246DOI Listing
February 2021

Transcutaneous near-infrared spectroscopy (NIRS) for monitoring kidney and liver allograft perfusion.

Int J Clin Pract 2021 Jan 20:e14034. Epub 2021 Jan 20.

Pediatric Urology, Department of Women's and Children's Health, University of Padua, Padua, Italy.

Background: The use of transcutaneous near-infrared spectroscopy (NIRS) for monitoring the perfusion of kidney and liver allografts has been proposed in the last years. This device might detect an early decrease in allograft oxygenation allowing prompt detection of vascular complications.

Methods: A systematic review of the literature about the use of transcutaneous NIRS in monitoring allograft perfusion was performed according to the PRISMA guidelines.

Results: The authors screened 1313 papers. The search yielded five pertinent articles. Three of them reported the experience of NIRS in kidney transplantation and the other two dealt with its use in liver transplantation, for a total of 55 paediatric patients and 121 adults. In the studies concerning kidney transplantation, NIRS measurements were significantly related to serum creatinine, estimated glomerular filtration rate (eGFR), urinary neutrophil gelatinase-associated lipocalin (u-NGAL), serum lactate, resistive index assessed by Doppler-ultrasonography and systolic blood pressure. The two studies dealing with liver transplantation found a significant decrease in liver regional oxygenation, assessed by NIRS, before the occurrence of vascular complications.

Conclusions: Preliminary studies have related NIRS monitoring to kidney and liver allograft perfusion, both in adults and children. Further investigation is needed to establish the normal range of NIRS values and the factors influencing NIRS monitoring.
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http://dx.doi.org/10.1111/ijcp.14034DOI Listing
January 2021

Employing a systematic approach to biobanking and analyzing clinical and genetic data for advancing COVID-19 research.

Eur J Hum Genet 2021 Jan 17. Epub 2021 Jan 17.

Medical Genetics, University of Siena, Siena, Italy.

Within the GEN-COVID Multicenter Study, biospecimens from more than 1000 SARS-CoV-2 positive individuals have thus far been collected in the GEN-COVID Biobank (GCB). Sample types include whole blood, plasma, serum, leukocytes, and DNA. The GCB links samples to detailed clinical data available in the GEN-COVID Patient Registry (GCPR). It includes hospitalized patients (74.25%), broken down into intubated, treated by CPAP-biPAP, treated with O supplementation, and without respiratory support (9.5%, 18.4%, 31.55% and 14.8, respectively); and non-hospitalized subjects (25.75%), either pauci- or asymptomatic. More than 150 clinical patient-level data fields have been collected and binarized for further statistics according to the organs/systems primarily affected by COVID-19: heart, liver, pancreas, kidney, chemosensors, innate or adaptive immunity, and clotting system. Hierarchical clustering analysis identified five main clinical categories: (1) severe multisystemic failure with either thromboembolic or pancreatic variant; (2) cytokine storm type, either severe with liver involvement or moderate; (3) moderate heart type, either with or without liver damage; (4) moderate multisystemic involvement, either with or without liver damage; (5) mild, either with or without hyposmia. GCB and GCPR are further linked to the GCGDR, which includes data from whole-exome sequencing and high-density SNP genotyping. The data are available for sharing through the Network for Italian Genomes, found within the COVID-19 dedicated section. The study objective is to systematize this comprehensive data collection and begin identifying multi-organ involvement in COVID-19, defining genetic parameters for infection susceptibility within the population, and mapping genetically COVID-19 severity and clinical complexity among patients.
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http://dx.doi.org/10.1038/s41431-020-00793-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811682PMC
January 2021

Impact of COVID-19 Pandemic in Children with CKD or Immunosuppression.

Clin J Am Soc Nephrol 2020 Dec 14. Epub 2020 Dec 14.

Pediatric Nephrology, Dialysis and Transplant Unit, Fondazione Istituto di Ricerca e Cura a Carattere Scientifico Cà Granda, Ospedale Maggiore Policlinico, Milan, Italy

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http://dx.doi.org/10.2215/CJN.13120820DOI Listing
December 2020

Variants in the SK2 channel gene (KCNN2) lead to dominant neurodevelopmental movement disorders.

Brain 2020 12;143(12):3564-3573

Institut du Cerveau et de la Moelle épinière (ICM), Sorbonne Université, UMR S 1127, Inserm U1127, CNRS UMR 7225, F-75013 Paris, France.

KCNN2 encodes the small conductance calcium-activated potassium channel 2 (SK2). Rodent models with spontaneous Kcnn2 mutations show abnormal gait and locomotor activity, tremor and memory deficits, but human disorders related to KCNN2 variants are largely unknown. Using exome sequencing, we identified a de novo KCNN2 frameshift deletion in a patient with learning disabilities, cerebellar ataxia and white matter abnormalities on brain MRI. This discovery prompted us to collect data from nine additional patients with de novo KCNN2 variants (one nonsense, one splice site, six missense variants and one in-frame deletion) and one family with a missense variant inherited from the affected mother. We investigated the functional impact of six selected variants on SK2 channel function using the patch-clamp technique. All variants tested but one, which was reclassified to uncertain significance, led to a loss-of-function of SK2 channels. Patients with KCNN2 variants had motor and language developmental delay, intellectual disability often associated with early-onset movement disorders comprising cerebellar ataxia and/or extrapyramidal symptoms. Altogether, our findings provide evidence that heterozygous variants, likely causing a haploinsufficiency of the KCNN2 gene, lead to novel autosomal dominant neurodevelopmental movement disorders mirroring phenotypes previously described in rodents.
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http://dx.doi.org/10.1093/brain/awaa346DOI Listing
December 2020

Clinical and molecular characterization of COVID-19 hospitalized patients.

PLoS One 2020 18;15(11):e0242534. Epub 2020 Nov 18.

Medical Genetics, University of Siena, Siena, Italy.

Clinical and molecular characterization by Whole Exome Sequencing (WES) is reported in 35 COVID-19 patients attending the University Hospital in Siena, Italy, from April 7 to May 7, 2020. Eighty percent of patients required respiratory assistance, half of them being on mechanical ventilation. Fiftyone percent had hepatic involvement and hyposmia was ascertained in 3 patients. Searching for common genes by collapsing methods against 150 WES of controls of the Italian population failed to give straightforward statistically significant results with the exception of two genes. This result is not unexpected since we are facing the most challenging common disorder triggered by environmental factors with a strong underlying heritability (50%). The lesson learned from Autism-Spectrum-Disorders prompted us to re-analyse the cohort treating each patient as an independent case, following a Mendelian-like model. We identified for each patient an average of 2.5 pathogenic mutations involved in virus infection susceptibility and pinpointing to one or more rare disorder(s). To our knowledge, this is the first report on WES and COVID-19. Our results suggest a combined model for COVID-19 susceptibility with a number of common susceptibility genes which represent the favorite background in which additional host private mutations may determine disease progression.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0242534PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7673557PMC
December 2020

Results of the PROPINE randomized controlled study suggest tapering of prednisone treatment for relapses of steroid sensitive nephrotic syndrome is not necessary in children.

Kidney Int 2021 02 2;99(2):475-483. Epub 2020 Nov 2.

Division of Nephrology and Dialysis, Department of Pediatric Subspecialties, Bambino Gesù Children's Hospital-IRCCS, Rome, Italy. Electronic address:

Corticosteroid-related toxicity in children with steroid-sensitive nephrotic syndrome is primarily related to the cumulative dose of prednisone. To optimize treatment of relapses, we conducted the PROPINE study, a multicentric, open-label, randomized, superiority trial. Seventy-eight relapsing children aged 3-17 years who had not received steroid-sparing medications during the previous 12 months were randomized to receive, from day five after remission, either 18 doses of 40 mg/m of prednisone on alternate days (short arm), or the same cumulative dose tapered over double the time (long arm). Patients were monitored with an ad-hoc smartphone application, allowing daily reporting. The primary outcome was the six-month relapse rate at which time, 23/40 and 16/38 patients had relapsed in the long and short arms, respectively (no significant difference). Additionally, 40/78 patients were also enrolled in a secondary crossover study and were allocated to the opposite arm. Altogether, at six months, the relapse rate was 32/40 and 28/40 in the long and short arms, respectively (no significant difference). A post-hoc analysis excluding 30 patients treated with low-dose prednisone maintenance therapy failed to show significant differences between the two arms. No differences in adverse events, blood pressure and weight gain were observed. Thus, our data do not support the prescription of prolonged tapering schedules for relapses of steroid-sensitive nephrotic syndrome in children.
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http://dx.doi.org/10.1016/j.kint.2020.09.024DOI Listing
February 2021

Pediatric transplantation in Europe during the COVID-19 pandemic: Early impact on activity and healthcare.

Clin Transplant 2020 10 22;34(10):e14063. Epub 2020 Sep 22.

La Paz Institute of Biomedical Research - IdiPAZ, La Paz University Hospital, Madrid, Spain.

The current pandemic SARS-CoV-2 has required an unusual allocation of resources that can negatively impact chronically ill patients and high-complexity procedures. Across the European Reference Network on Pediatric Transplantation (ERN TransplantChild), we conducted a survey to investigate the impact of the COVID-19 outbreak on pediatric transplant activity and healthcare practices in both solid organ transplantation (SOT) and hematopoietic stem cell transplantation (HSCT). The replies of 30 professionals from 18 centers in Europe were collected. Twelve of 18 centers (67%) showed a reduction in their usual transplant activity. Additionally, outpatient visits have been modified and restricted to selected ones, and the use of telemedicine tools has increased. Additionally, a total of 14 COVID-19 pediatric transplanted patients were identified at the time of the survey, including eight transplant recipients and six candidates for transplantation. Only two moderate-severe cases were reported, both in HSCT setting. These survey results demonstrate the limitations in healthcare resources for pediatric transplantation patients during early stages of this pandemic. COVID-19 disease is a major worldwide challenge for the field of pediatric transplantation, where there will be a need for systematic data collection, encouraging regular discussions to address the long-term consequences for pediatric transplantation candidates, recipients, and their families.
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http://dx.doi.org/10.1111/ctr.14063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7435500PMC
October 2020

ACE2 gene variants may underlie interindividual variability and susceptibility to COVID-19 in the Italian population.

Eur J Hum Genet 2020 11 17;28(11):1602-1614. Epub 2020 Jul 17.

Genetica Medica, Azienda Ospedaliera Universitaria Senese, Siena, Italy.

In December 2019, an initial cluster of interstitial bilateral pneumonia emerged in Wuhan, China. A human-to-human transmission was assumed and a previously unrecognized entity, termed coronavirus disease-19 (COVID-19) due to a novel coronavirus (SARS-CoV-2) was described. The infection has rapidly spread out all over the world and Italy has been the first European country experiencing the endemic wave with unexpected clinical severity in comparison with Asian countries. It has been shown that SARS-CoV-2 utilizes angiotensin converting enzyme 2 (ACE2) as host receptor and host proteases for cell surface binding and internalization. Thus, a predisposing genetic background can give reason for interindividual disease susceptibility and/or severity. Taking advantage of the Network of Italian Genomes (NIG), here we mined whole-exome sequencing data of 6930 Italian control individuals from five different centers looking for ACE2 variants. A number of variants with a potential impact on protein stability were identified. Among these, three more common missense changes, p.(Asn720Asp), p.(Lys26Arg), and p.(Gly211Arg) were predicted to interfere with protein structure and stabilization. Rare variants likely interfering with the internalization process, namely p.(Leu351Val) and p.(Pro389His), predicted to interfere with SARS-CoV-2 spike protein binding, were also observed. Comparison of ACE2 WES data between a cohort of 131 patients and 258 controls allowed identifying a statistically significant (P value < 0.029) higher allelic variability in controls compared with patients. These findings suggest that a predisposing genetic background may contribute to the observed interindividual clinical variability associated with COVID-19, allowing an evidence-based risk assessment leading to personalized preventive measures and therapeutic options.
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http://dx.doi.org/10.1038/s41431-020-0691-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7366459PMC
November 2020

AAV-mediated FOXG1 gene editing in human Rett primary cells.

Eur J Hum Genet 2020 10 15;28(10):1446-1458. Epub 2020 Jun 15.

Medical Genetics, University of Siena, Siena, Italy.

Variations in the Forkhead Box G1 (FOXG1) gene cause FOXG1 syndrome spectrum, including the congenital variant of Rett syndrome, characterized by early onset of regression, Rett-like and jerky movements, and cortical visual impairment. Due to the largely unknown pathophysiological mechanisms downstream the impairment of this transcriptional regulator, a specific treatment is not yet available. Since both haploinsufficiency and hyper-expression of FOXG1 cause diseases in humans, we reasoned that adding a gene under nonnative regulatory sequences would be a risky strategy as opposed to a genome editing approach where the mutated gene is reversed into wild-type. Here, we demonstrate that an adeno-associated viruses (AAVs)-coupled CRISPR/Cas9 system is able to target and correct FOXG1 variants in patient-derived fibroblasts, induced Pluripotent Stem Cells (iPSCs) and iPSC-derived neurons. Variant-specific single-guide RNAs (sgRNAs) and donor DNAs have been selected and cloned together with a mCherry/EGFP reporter system. Specific sgRNA recognition sequences were inserted upstream and downstream Cas9 CDS to allow self-cleavage and inactivation. We demonstrated that AAV serotypes vary in transduction efficiency depending on the target cell type, the best being AAV9 in fibroblasts and iPSC-derived neurons, and AAV2 in iPSCs. Next-generation sequencing (NGS) of mCherry/EGFP transfected cells demonstrated that the mutated alleles were repaired with high efficiency (20-35% reversion) and precision both in terms of allelic discrimination and off-target activity. The genome editing strategy tested in this study has proven to precisely repair FOXG1 and delivery through an AAV9-based system represents a step forward toward the development of a therapy for Rett syndrome.
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http://dx.doi.org/10.1038/s41431-020-0652-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608362PMC
October 2020

Sedentariness and Urinary Metabolite Profile in Type 2 Diabetic Patients, a Cross-Sectional Study.

Metabolites 2020 May 18;10(5). Epub 2020 May 18.

Department of Drug Science and Technology University of Turin, 10125 Turin, Italy.

Recent findings indicate a significant association between sedentary (SED)-time and type 2 diabetes mellitus(T2DM). The aim of this study was to investigate whether different levels of SED-time could impact on biochemical and physiological processes occurring in sedentary and physically inactive T2DM patients. In particular, patients from the "Italian Diabetes and Exercise Study (IDES)_2 trial belonging to the first and fourth quartile of SED-time were compared. Urine samples were analyzed by comprehensive two-dimensional gas chromatography(GC×GC) with parallel detection by mass spectrometry and flame ionization detection(GC×2GC-MS/FID). This platform enables accurate profiling and fingerprinting of urinary metabolites while maximizing the overall information capacity, quantitation reliability, and response linearity. Moreover, using advanced pattern recognition, the fingerprinting process was extended to untargeted and targeted features, revealing diagnostic urinary fingerprints between groups. Quantitative metabolomics was then applied to analytes of relevance for robust comparisons. Increased levels of glycine, L-valine,L-threonine, L-phenylalanine, L-leucine, L-alanine, succinic acid, 2-ketoglutaric acid, xylitol, and ribitol were revealed in samples from less sedentary women. In conclusion, SED-time is associated with changes in urine metabolome signatures. These preliminary results suggest that reducing SED-time could be a strategy to improve the health status of a large proportion of diabetic patients.
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http://dx.doi.org/10.3390/metabo10050205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7281751PMC
May 2020

High rate of HDR in gene editing of p.(Thr158Met) MECP2 mutational hotspot.

Eur J Hum Genet 2020 09 24;28(9):1231-1242. Epub 2020 Apr 24.

Medical Genetics, University of Siena, Siena, Italy.

Rett syndrome is a progressive neurodevelopmental disorder which affects almost exclusively girls, caused by variants in MECP2 gene. Effective therapies for this devastating disorder are not yet available and the need for tight regulation of MECP2 expression for brain to properly function makes gene replacement therapy risky. For this reason, gene editing with CRISPR/Cas9 technology appears as a preferable option for the development of new therapies. To study the disease, we developed and characterized a human neuronal model obtained by genetic reprogramming of patient-derived primary fibroblasts into induced Pluripotent Stem Cells. This cellular model represents an important source for our studies, aiming to correct MECP2 variants in neurons which represent the primarily affected cell type. We engineered a gene editing toolkit composed by a two-plasmid system to correct a hotspot missense variant in MECP2, c.473 C > T (p.(Thr158Met)). The first construct expresses the variant-specific sgRNA and the Donor DNA along with a fluorescent reporter system. The second construct brings Cas9 and targets for auto-cleaving, to avoid long-term Cas9 expression. NGS analysis on sorted cells from four independent patients demonstrated an exceptionally high editing efficiency, with up to 80% of HDR and less than 1% of indels in all patients, outlining the relevant potentiality of the approach for Rett syndrome therapy.
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http://dx.doi.org/10.1038/s41431-020-0624-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7609331PMC
September 2020

Functional Study of Novel Bartter's Syndrome Mutations in ClC-Kb and Rescue by the Accessory Subunit Barttin Toward Personalized Medicine.

Front Pharmacol 2020 17;11:327. Epub 2020 Mar 17.

Department of Pharmacy-Drug Sciences, University of Bari "Aldo Moro", Bari, Italy.

Type III and IV Bartter syndromes (BS) are rare kidney tubulopathies caused by loss-of-function mutations in the and genes coding respectively for the ClC-Kb chloride channels and accessory subunit barttin. ClC-K channels are expressed in the Henle's loop, distal convoluted tubule, and cortical collecting ducts of the kidney and contribute to chloride absorption and urine concentration. In our Italian cohort, we identified two new mutations in , G167V and G289R, in children affected by BS and previously reported genetic variants, A242E, a chimeric gene and the deletion of the whole . All the patients had hypokalemia and metabolic alkalosis, increased serum renin and aldosterone levels and were treated with a symptomatic therapy. In order to define the molecular mechanisms responsible for BS, we co-expressed ClC-Kb wild type and channels with point mutations with barttin in HEK 293 cells and characterized chloride currents through the patch-clamp technique. In addition, we attempted to revert the functional defect caused by BS mutations through barttin overexpression. G167V and A242E channels showed a drastic current reduction compared to wild type, likely suggesting compromised expression of mutant channels at the plasma membrane. Conversely, G289R channel was similar to wild type raising the doubt that an additional mutation in another gene or other mechanisms could account for the clinical phenotype. Interestingly, increasing ClC-K/barttin ratio augmented G167V and A242E mutants' chloride current amplitudes towards wild type levels. These results confirm a genotype-phenotype correlation in BS and represent a preliminary proof of concept that molecules functioning as molecular chaperones can restore channel function in expression-defective ClC-Kb mutants.
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http://dx.doi.org/10.3389/fphar.2020.00327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7092721PMC
March 2020

New frontiers to cure Alport syndrome: COL4A3 and COL4A5 gene editing in podocyte-lineage cells.

Eur J Hum Genet 2020 04 21;28(4):480-490. Epub 2019 Nov 21.

Genetica Medica, Azienda Ospedaliera Universitaria Senese, Siena, Italy.

Alport syndrome (AS) is an inherited genetic disorder characterized by range of alterations from glomerular basement membrane abnormalities up to end-stage renal disease. Pathogenic variants in the collagen α3, α4, and α5 encoding genes are causative both of the autosomal dominant and of the X-linked forms of AS. Podocytes are the only renal cells that are able to produce the COL(IV)a3-a4a5 heterotrimer. We have previously demonstrated how it is possible to isolate podocyte-lineage cells from urine of patients, providing an easily accessible cellular model closer to the podocytes' physiological conditions. Taking advantage of disease-relevant cell lines, we employed a two-plasmid approach in order to achieve a beneficial and stable variant-specific correction using CRISPR/Cas9 genome editing. One plasmid carries a Donor DNA and a reporter system mCherry/GFP to track the activity of Cas9 in cells. The other plasmid carries a self-cleaving SpCas9 and the variant-specific sgRNA. We have analyzed two stable podocyte-lineage cell lines, harboring a variant in the X-linked COL4A5 (p.(Gly624Asp)) and in the autosomal COL4A3 gene (p.(Gly856Glu)). We have achieved reversion of variants greater than 40% with undesired insertions/deletions lower than 15%. Overall, we have demonstrated a new gene therapy approach directly on patients' cells, key players of Alport pathogenesis, and we have reverted COL4 causative variants towards the wild type state. These results, in combination with preclinical models, could open new frontiers in the management and the treatment of the disorder.
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http://dx.doi.org/10.1038/s41431-019-0537-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7080842PMC
April 2020

Post-transplant recurrence of steroid resistant nephrotic syndrome in children: the Italian experience.

J Nephrol 2020 Aug 15;33(4):849-857. Epub 2019 Oct 15.

Pediatric Nephrology, Dialysis and Transplant Unit, Department of Clinical Sciences and Community Health, Fondazione IRCCS Ca' Granda-Ospedale Maggiore Policlinico, University of Milan, via della Commenda, 9, 20122, Milan, Italy.

Background: Steroid resistant nephrotic syndrome (SRNS) is a frequent cause of end stage renal disease in children and post-transplant disease recurrence is a major cause of graft loss.

Methods: We identified all children with SRNS who underwent renal transplantation in Italy, between 2005 and 2017. Data were retrospectively collected for the presence of a causative gene mutation, sex, histology, duration of pre-transplant dialysis, age at onset and transplant, HLA matching, recurrence, therapy for recurrence, and graft survival.

Results: 101 patients underwent a first and 22 a second renal transplant. After a median follow-up of 58.5 months, the disease recurred on the first renal transplant in 53.3% of patients with a non-genetic and none with a genetic SRNS. Age at transplant > 9 years and the presence of at least one HLA-AB match were independent risk factors for recurrence. Duration of dialysis was longer in children with relapse, but did not reach statistical significance. Overall, 24% of patients lost the first graft, with recurrence representing the commonest cause. Among 22 patients who underwent a second transplant, 5 suffered of SRNS recurrence. SRNS relapsed in 5/9 (55%) patients with disease recurrence in their first transplant and 2 of them lost the second graft.

Conclusions: Absence of a causative mutation represents the major risk factor for post-transplant recurrence in children with SRNS, while transplant can be curative in genetic SRNS. A prolonged time spent on dialysis before transplantation has no protective effect on the risk of relapse and should not be encouraged. Retransplantation represents a second chance after graft loss for recurrence.
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http://dx.doi.org/10.1007/s40620-019-00660-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7381476PMC
August 2020

Paclitaxel-Loaded Nanosponges Inhibit Growth and Angiogenesis in Melanoma Cell Models.

Front Pharmacol 2019 12;10:776. Epub 2019 Jul 12.

Dipartimento di Scienza e Tecnologia del Farmaco, University of Torino, Torino, Italy.

This study investigated the effects of free paclitaxel (PTX) and PTX-loaded in pyromellitic nanosponges (PTX-PNS) in reducing and melanoma cell growth and invasivity, and in inhibiting angiogenesis. To test the response of cells to the two PTX formulations, the cell viability was evaluated by MTT assay in seven continuous cell lines, in primary melanoma cells, both in 2D and 3D cultures, and in human umbilical vein endothelial cells (HUVECs) after exposure to different concentrations of PTX or PTX-PNS. Cell motility was assessed by a scratch assay or Boyden chamber assay, evaluating cell migration in presence or absence of diverse concentrations of PTX or PTX-PNS. The effect of PTX and PTX-PNS on angiogenesis was evaluated as endothelial tube formation assay, a test able to estimate the formation of three-dimensional vessels . To assess the anticancer effect of PTX and PTX-PNS in experiments, the two drug formulations were tested in a melanoma mouse model obtained by B16-BL6 cell implantation in C57/BL6 mice. Results obtained were as follows: 1) MTT analysis revealed that cell proliferation was more affected by PTX-PNS than by PTX in all tested cell lines, in both 2D and 3D cultures; 2) the analysis of the cell migration showed that PTX-PNS acted at very lower concentrations than PTX; 3) tube formation assay showed that PTX-PNS were more effective in inhibiting tube formation than free PTX; and 4) experiments demonstrated that tumor weights, volumes, and growth were significantly reduced by PTX-PNS treatment with respect to PTX; the angiogenesis and the cell proliferation, detected in the tumor samples with CD31 and Ki-67 antibodies, respectively, indicated that, in the PTX-PNS-treated tumors, the tube formation was inhibited, and a low amount of proliferating cells was present. Taken together, our data demonstrated that our new PTX nanoformulation can respond to some important issues related to PTX treatment, lowering the anti-tumor effective doses and increasing the effectiveness in inhibiting melanoma growth .
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http://dx.doi.org/10.3389/fphar.2019.00776DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6639435PMC
July 2019

Effect of Bilastine on Diabetic Nephropathy in DBA2/J Mice.

Int J Mol Sci 2019 May 24;20(10). Epub 2019 May 24.

Department of Scienza e Tecnologia del Farmaco, University of Turin, Via P. Giuria 9, 10125 Turin, Italy.

Diabetic nephropathy is an unmet therapeutic need, and the search for new therapeutic strategies is warranted. Previous data point to histamine H receptor as a possible target for glomerular dysfunction associated with long term hyperglycaemia. Therefore, this study investigated the effects of the H receptor antagonist bilastine on renal morphology and function in a murine model of streptozotocin-induced diabetes. Diabetes was induced in DBA2/J male mice and, from diabetes onset (glycaemia ≥200 mg/dL), mice received bilastine (1-30 mg/kg/day) by oral gavage for 14 consecutive weeks. At the end of the experimental protocol, diabetic mice showed polyuria (+195.5%), increase in Albumin-to-Creatine Ratio (ACR, +284.7%), and a significant drop in creatinine clearance ( < 0.05). Bilastine prevented ACR increase and restored creatinine clearance in a dose-dependent manner, suggesting a positive effect on glomerular filtration. The ultrastructural analysis showed a preserved junctional integrity. Preservation of the basal nephrin, P-cadherin, and synaptopodin expression could explain this effect. In conclusion, the H receptor could contribute to the glomerular damage occurring in diabetic nephropathy. Bilastine preserved the glomerular junctional integrity, leading to the hypothesis of anti-H antihistamines as a possible add-on therapy for diabetic nephropathy.
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http://dx.doi.org/10.3390/ijms20102554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6566437PMC
May 2019

Analyzing Cysteine Site Neighbors in Proteins to Reveal Dimethyl Fumarate Targets.

Proteomics 2019 02 25;19(4):e1800301. Epub 2019 Jan 25.

Dipartimento di Scienza e Tecnologia del Farmaco, Università degli Studi di Torino, Turin, 10125, Italy.

This work proposes a novel approach by which to consistently classify cysteine sites in proteins in terms of their reactivity toward dimethyl fumarate (DMF) and fumarate. Dimethyl fumarate-based drug products have been approved for use as oral treatments for psoriasis and relapsing-remitting multiple sclerosis. The adduction of DMF and its (re)active metabolites to certain cysteine residues in proteins is thought to underlie their effects. However, only a few receptors for these compounds have been discovered to date. Our approach takes advantage of the growing number of known DMF- and fumarate-sensitive proteins and sites to perform analyses by combining the concepts of network theory, for protein structure analyses, and machine-learning procedures. Wide-ranging and previously unforeseen variety is found in the analysis of the neighborhood composition (the first neighbors) of cysteine sites found in DMF- and fumarate-sensitive proteins. Furthermore, neighborhood composition has shown itself to be a network-type attribute that is endowed with remarkable predictive power when distinct classification algorithms are employed. In conclusion, when adopted in combination with other target identification/validation approaches, methods that are based on the analysis of cysteine site neighbors in proteins should provide useful information by which to decipher the mode of action of DMF-based drugs.
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http://dx.doi.org/10.1002/pmic.201800301DOI Listing
February 2019

Could the interaction between LMX1B and PAX2 influence the severity of renal symptoms?

Eur J Hum Genet 2018 11 4;26(11):1708-1712. Epub 2018 Jul 4.

Laboratory of Immunopathology and Molecular Biology of the Kidney, Department of Women's and Children's Health, University of Padova, Padua, Italy.

Nail Patella syndrome (NPS) is a rare autosomal dominant disease characterized by varying degrees of patella, nail, and elbows dysplasia and also ocular and renal congenital abnormalities. The renal involvement, ranging from hematuria and proteinuria to end-stage renal disease, is present in 22-60% of NPS cases. Heterozygous variants in LMX1B are known to be responsible of NPS and it has been hypothesized that the variable expressivity is due to the interaction of LMX1B with other developmental genes. We reported a case of co-presence of LMX1B and PAX2 variants in a child with extrarenal manifestation of NPS and end-stage renal disease but congenital bilateral renal hypodysplasia and vesicoureteral reflux. The LMX1B variant was de novo, whereas the PAX2 variant was inherited from the mother that had bilateral renal hypoplasia although in presence of only a mild chronic kidney disease. The molecular interaction between LMX1B and PAX2 has been already reported in vitro and this finding suggest that the worst renal NPS phenotype of our patient could be due to the defective expression of these two genes during nephrogenesis. In conclusion, our finding suggests that PAX2 may act as modifier gene in Nail Patella phenotype.
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http://dx.doi.org/10.1038/s41431-018-0213-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6189091PMC
November 2018

Effects of vitamin D on insulin resistance and myosteatosis in diet-induced obese mice.

PLoS One 2018 17;13(1):e0189707. Epub 2018 Jan 17.

Division of Endocrinology, Diabetology and Metabolism, Department of Medical Sciences, University of Turin, Turin, Italy.

Epidemiological studies pointed out to a strong association between vitamin D deficiency and type 2 diabetes prevalence. However, the role of vitamin D supplementation in the skeletal muscle, a tissue that play a crucial role in the maintenance of glucose homeostasis, has been scarcely investigated so far. On this basis, this study aimed to evaluate the effect of vitamin D supplementation in a murine model of diet-induced insulin resistance with particular attention to the effects evoked on the skeletal muscle. Male C57BL/6J mice (n = 40) were fed with a control or a High Fat-High Sugar (HFHS) diet for 4 months. Subsets of animals were treated for 2 months with vitamin D (7 μg·kg-1, i.p. three times/week). HFHS diet induced body weight increase, hyperglycemia and impaired glucose tolerance. HFHS animals showed an impaired insulin signaling and a marked fat accumulation in the skeletal muscle. Vitamin D reduced body weight and improved systemic glucose tolerance. In addition, vitamin D restored the impaired muscle insulin signaling and reverted myosteatosis evoked by the diet. These effects were associated to decreased activation of NF-κB and lower levels of TNF-alpha. Consistently, a significantly decreased activation of the SCAP/SREBP lipogenic pathway and lower levels of CML protein adducts and RAGE expression were observed in skeletal muscle of animals treated with vitamin D. Collectively, these data indicate that vitamin D-induced selective inhibition of signaling pathways (including NF-κB, SCAP/SREBP and CML/RAGE cascades) within the skeletal muscle significantly contributed to the beneficial effects of vitamin D supplementation against diet-induced metabolic derangements.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0189707PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5771572PMC
January 2018

Usefulness of Tc-dimercaptosuccinic acid renal scan in the diagnosis and follow-up of acute tubulointerstitial nephritis in children.

Clin Kidney J 2017 Oct 12;10(5):655-660. Epub 2017 Jun 12.

Nephrology, Dialysis and Transplant Unit, Department of Women's and Children's Health, University-Hospital of Padova, Padova, Italy.

Background: Symptoms and signs of acute tubulointerstitial nephritis (ATIN) are nonspecific; therefore, renal biopsy is often necessary to clarify the diagnosis. The aim of this study was to evaluate the use of Tc-dimercaptosuccinic acid (DMSA) scintigraphy in the diagnosis and follow-up of ATIN.

Methods: We retrospectively reviewed the charts of five patients (nine renal units) with a median age of 14 years who underwent DMSA scan after a clinical and/or biopsy-proven diagnosis of ATIN. The exam was performed within 1 month after disease onset and repeated at a median time of 12 months after the acute phase.

Results: DMSA renal scans performed during the acute phase allowed the discovery of suggestive findings, including diffuse reduction of the renal uptake of radionuclide and presence of multiple 'cold' focal lesions in a corticomedullary distribution. The follow-up scintigraphy resulted normal in two patients who were treated with steroids and in one patient who presented a mild renal dysfunction in the acute phase. By contrast, the control scan showed persistent renal damage in one patient who was further readmitted because of hypertension and in one renal transplanted patient who presented a Stage 3 acute kidney injury in the acute phase.

Conclusions: DMSA renal scan might be a reliable tool for an early non-invasive diagnosis of ATIN in children and might be particularly useful in those patients who are not candidates for a kidney biopsy. Moreover, DMSA scan gives accurate follow-up evaluation, as it allows monitoring of the evolution of acute renal parenchymal inflammation with potential risk of renal scar formation. Due to the small sample size, our findings warrant further validation in a larger study.
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http://dx.doi.org/10.1093/ckj/sfx041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5622892PMC
October 2017

The Italian Society for Pediatric Nephrology (SINePe) consensus document on the management of nephrotic syndrome in children: Part I - Diagnosis and treatment of the first episode and the first relapse.

Ital J Pediatr 2017 Apr 21;43(1):41. Epub 2017 Apr 21.

Institute of Maternal and Child Health IRCCS "Burlo Garofolo", Department of Pediatrics, Trieste, Italy.

This consensus document is aimed at providing an updated, multidisciplinary overview on the diagnosis and treatment of pediatric nephrotic syndrome (NS) at first presentation. It is the first consensus document of its kind to be produced by all the pediatric nephrology centres in Italy, in line with what is already present in other countries such as France, Germany and the USA. It is based on the current knowledge surrounding the symptomatic and steroid treatment of NS, with a view to providing the basis for a separate consensus document on the treatment of relapses. NS is one of the most common pediatric glomerular diseases, with an incidence of around 2-7 cases per 100000 children per year. Corticosteroids are the mainstay of treatment, but the optimal therapeutic regimen for managing childhood idiopathic NS is still under debate. In Italy, shared treatment guidelines were lacking and, consequently, the choice of steroid regimen was based on the clinical expertise of each individual unit. On the basis of the 2015 Cochrane systematic review, KDIGO Guidelines and more recent data from the literature, this working group, with the contribution of all the pediatric nephrology centres in Italy and on the behalf of the Italian Society of Pediatric Nephrology, has produced a shared steroid protocol that will be useful for National Health System hospitals and pediatricians. Investigations at initial presentation and the principal causes of NS to be screened are suggested. In the early phase of the disease, symptomatic treatment is also important as many severe complications can occur which are either directly related to the pathophysiology of the underlying NS or to the steroid treatment itself. To date, very few studies have been published on the prophylaxis and treatment of these early complications, while recommendations are either lacking or conflicting. This consensus provides indications for the prevention, early recognition and treatment of these complications (management of edema and hypovolemia, therapy and prophylaxis of infections and thromboembolic events). Finally, recommendations about the clinical definition of steroid resistance and its initial diagnostic management, as well as indications for renal biopsy are provided.
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http://dx.doi.org/10.1186/s13052-017-0356-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5399429PMC
April 2017

Microvesicles released from fat-laden cells promote activation of hepatocellular NLRP3 inflammasome: A pro-inflammatory link between lipotoxicity and non-alcoholic steatohepatitis.

PLoS One 2017 1;12(3):e0172575. Epub 2017 Mar 1.

Department of Clinical and Biological Sciences, University of Torino, Torino, Italy.

Non-Alcoholic Fatty Liver Disease (NAFLD) is a major form of chronic liver disease in the general population in relation to its high prevalence among overweight/obese individuals and patients with diabetes type II or metabolic syndrome. NAFLD can progress to steatohepatitis (NASH), fibrosis and cirrhosis and end-stage of liver disease but mechanisms involved are still incompletely characterized. Within the mechanisms proposed to mediate the progression of NAFLD, lipotoxicity is believed to play a major role. In the present study we provide data suggesting that microvesicles (MVs) released by fat-laden cells undergoing lipotoxicity can activate NLRP3 inflammasome following internalization by either cells of hepatocellular origin or macrophages. Inflammasome activation involves NF-kB-mediated up-regulation of NLRP3, pro-caspase-1 and pro-Interleukin-1, then inflammasome complex formation and Caspase-1 activation leading finally to an increased release of IL-1β. Since the release of MVs from lipotoxic cells and the activation of NLRP3 inflammasome have been reported to occur in vivo in either clinical or experimental NASH, these data suggest a novel rational link between lipotoxicity and increased inflammatory response.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0172575PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5331985PMC
August 2017

The genetic and clinical spectrum of a large cohort of patients with distal renal tubular acidosis.

Kidney Int 2017 05 21;91(5):1243-1255. Epub 2017 Feb 21.

Department of Biomedical Experimental and Clinical Sciences "Mario Serio", University of Florence, Florence, Italy; Medical Genetics Unit, Meyer Children's University Hospital, Florence, Italy. Electronic address:

Primary distal renal tubular acidosis is a rare genetic disease. Mutations in SLC4A1, ATP6V0A4, and ATP6V1B1 genes have been described as the cause of the disease, transmitted as either an autosomal dominant or recessive trait. Particular clinical features, such as sensorineural hearing loss, have been mainly described in association with mutations in one gene instead of the others. Nevertheless, the diagnosis of distal renal tubular acidosis is essentially based on clinical and laboratory findings, and the series of patients described so far are usually represented by small cohorts. Therefore, a strict genotype-phenotype correlation is still lacking, and questions about whether clinical and laboratory data should direct the genetic analysis remain open. Here, we applied next-generation sequencing in 89 patients with a clinical diagnosis of distal renal tubular acidosis, analyzing the prevalence of genetic defects in SLC4A1, ATP6V0A4, and ATP6V1B1 genes and the clinical phenotype. A genetic cause was determined in 71.9% of cases. In our group of sporadic cases, clinical features, including sensorineural hearing loss, are not specific indicators of the causal underlying gene. Mutations in the ATP6V0A4 gene are quite as frequent as mutations in ATP6V1B1 in patients with recessive disease. Chronic kidney disease was frequent in patients with a long history of the disease. Thus, our results suggest that when distal renal tubular acidosis is suspected, complete genetic testing could be considered, irrespective of the clinical phenotype of the patient.
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http://dx.doi.org/10.1016/j.kint.2016.12.017DOI Listing
May 2017

Effects of linagliptin on human immortalized podocytes: a cellular system to study dipeptidyl-peptidase 4 inhibition.

Br J Pharmacol 2017 05 6;174(9):809-821. Epub 2017 Mar 6.

Dipartimento di Scienza e Tecnologia del Farmaco, Università degli Studi di Torino, Turin, Italy.

Background And Purpose: Dipeptidyl-peptidase 4 (DPP4) is expressed by resident renal cells, including glomerular cells. DPP4 inhibitors (gliptins) exert albuminuria lowering effects, but the role of renal DPP4 as a pharmacological target has not been elucidated. To better understand the actions of gliptins, the effects of linagliptin on the behaviour of immortalized human podocytes and mesangial cells were evaluated.

Experimental Approach: The expression of DPP4 was measured at both the mRNA and protein levels. The effects of linagliptin on DPP4 activity, cell growth and cell cycle progression were determined. The contribution of the stromal cell-derived factor-1- CXCR4/CXCR7 signalling pathways was evaluated by studying the effects of AMD3100 (a CXCR4 antagonist and CXCR7 agonist) alone and in combination with linagliptin. The contribution of ERK1/2 activation was analysed by studying the effects of the MAPK kinase 1/2 inhibitor AZD6244.

Key Results: DPP4 was highly expressed in podocytes. The activity of DPP4 and podocyte growth were reduced by linagliptin. The effects of sitagliptin on podocyte growth were similar to those of linagliptin, were associated with inhibition of cell proliferation and mimicked by AMD3100. Moreover, linagliptin and AMD3100 were found to have a synergistic interaction, whereas no interaction was seen between linagliptin and AZD6244.

Conclusions And Implications: Our cultures of human glomerular cells represent a reliable system for investigating the actions of gliptins. Moreover, DPP4 contributes to the regulation of podocyte behaviour. Inhibition of DPP4 in podocytes could underlie the effects of linagliptin on glomerular cells.
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http://dx.doi.org/10.1111/bph.13739DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5386990PMC
May 2017

Kidney Versus Combined Kidney and Liver Transplantation in Young People With Autosomal Recessive Polycystic Kidney Disease: Data From the European Society for Pediatric Nephrology/European Renal Association-European Dialysis and Transplant (ESPN/ERA-EDTA) Registry.

Am J Kidney Dis 2016 Nov 21;68(5):782-788. Epub 2016 Aug 21.

Department of Pediatric, Academic Medical Center, Amsterdam, the Netherlands.

Background: The choice for either kidney or combined liver-kidney transplantation in young people with kidney failure and liver fibrosis due to autosomal recessive polycystic kidney disease (ARPKD) can be challenging. We aimed to analyze the characteristics and outcomes of transplantation type in these children, adolescents, and young adults.

Study Design: Cohort study.

Setting & Participants: We derived data for children, adolescents, and young adults with ARPKD with either kidney or combined liver-kidney transplants for 1995 to 2012 from the ESPN/ERA-EDTA Registry, a European pediatric renal registry collecting data from 36 European countries.

Factor: Liver transplantation.

Outcomes & Measurements: Transplantation and patient survival.

Results: 202 patients with ARPKD aged 19 years or younger underwent transplantation after a median of 0.4 (IQR, 0.0-1.4) years on dialysis therapy at a median age of 9.0 (IQR, 4.1-13.7) years. 32 (15.8%) underwent combined liver-kidney transplantation, 163 (80.7%) underwent kidney transplantation, and 7 (3.5%) were excluded because transplantation type was unknown. Age- and sex-adjusted 5-year patient survival posttransplantation was 95.5% (95% CI, 92.4%-98.8%) overall: 97.4% (95% CI, 94.9%-100.0%) for patients with kidney transplantation in contrast to 87.0% (95% CI, 75.8%-99.8%) with combined liver-kidney transplantation. The age- and sex-adjusted risk for death after combined liver-kidney transplantation was 6.7-fold (95% CI, 1.8- to 25.4-fold) greater than after kidney transplantation (P=0.005). Five-year death-censored kidney transplant survival following combined liver-kidney and kidney transplantation was similar (92.1% vs 85.9%; P=0.4).

Limitations: No data for liver disease of kidney therapy recipients.

Conclusions: Combined liver-kidney transplantation in ARPKD is associated with increased mortality compared to kidney transplantation in our large observational study and was not associated with improved 5-year kidney transplant survival. Long-term follow-up of both kidney and liver involvement are needed to better delineate the optimal transplantation strategy.
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http://dx.doi.org/10.1053/j.ajkd.2016.06.019DOI Listing
November 2016

Renal transplantation in sensitized children and young adults: a nationwide approach.

Nephrol Dial Transplant 2017 01;32(1):191-195

Italian National Transplant Centre, Italian National Institute of Health (ISS), Rome, Italy.

Background: High levels of preformed anti-HLA antibodies dramatically diminish renal transplant outcomes. Most desensitization programmes guarantee good intermediate outcomes but quite disappointing long-term prognosis. The search for a fully compatible kidney increases time on the waiting list.

Methods: In February 2011, a nationwide hyperimmune programme (NHP) was begun in Italy: all available kidneys are primarily proposed to highly sensitized patients with a panel reactive antibody above 80%. In this manuscript, we evaluate the outcome of paediatric patients transplanted with this approach.

Results: Twenty-one patients were transplanted. Complete data are available for 20 patients. Mean age at transplantation was 14.5 years [standard deviation (SD) ± 5.5)]. Mean time on the waiting list was 29.3 months (SD ± 27.5). Median follow-up was 29.2 months (range: 11.2-59.3). The average number of HLA mismatches in these patients was 2.3 versus 3.7 in 48 standard patients transplanted in the same period (P < 0.001). Only one graft was lost. Two cases of humoral rejection occurred and were successfully treated. No cellular rejection was reported. Median creatinine clearance was 84, 88, 77 and 77 mL/min/1.73 m 2 respectively 1, 6, 12 and 24 months after transplant.

Conclusions: Transplantation of sensitized patients avoiding prohibited antigens is feasible, at least in a selected cohort of patients. In order to be able to further improve this approach, which in our opinion is very successful, it would be necessary to expand the donor pool, possibly increasing the number of countries participating in the programme. In this series, time on the waiting list did not increase significantly. This allocation policy should ideally lead to an outcome comparable to that expected in standard patients, which is particularly desirable in young patients who have the longest life expectancy. Since long-term results of desensitization programmes are not (yet) convincing, we suggest that these programmes should be reserved for selected cases where compatible organs cannot be found within a reasonable time span.
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http://dx.doi.org/10.1093/ndt/gfw369DOI Listing
January 2017

Deciphering Variability of PKD1 and PKD2 in an Italian Cohort of 643 Patients with Autosomal Dominant Polycystic Kidney Disease (ADPKD).

Sci Rep 2016 08 8;6:30850. Epub 2016 Aug 8.

IRCCS San Raffaele Scientific Institute, Division of Genetics and Cell Biology, Unit of Genomics for Human Disease Diagnosis, Milan, Italy.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most common hereditary kidney disease. We analysed PKD1 and PKD2, in a large cohort of 440 unrelated Italian patients with ADPKD and 203 relatives by direct sequencing and MLPA. Molecular and detailed phenotypic data have been collected and submitted to the PKD1/PKD2 LOVD database. This is the first large retrospective study in Italian patients, describing 701 variants, 249 (35.5%) already associated with ADPKD and 452 (64.5%) novel. According to the criteria adopted, the overall detection rate was 80% (352/440). Novel variants with uncertain significance were found in 14% of patients. Among patients with pathogenic variants, in 301 (85.5%) the disease is associated with PKD1, 196 (55.7%) truncating, 81 (23%) non truncating, 24 (6.8%) IF indels, and in 51 (14.5%) with PKD2. Our results outline the high allelic heterogeneity of variants, complicated by the presence of variants of uncertain significance as well as of multiple variants in the same subject. Classification of novel variants may be particularly cumbersome having an important impact on the genetic counselling. Our study confirms the importance to improve the assessment of variant pathogenicity for ADPKD; to this point databasing of both clinical and molecular data is crucial.
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http://dx.doi.org/10.1038/srep30850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4976333PMC
August 2016

Empagliflozin Protects against Diet-Induced NLRP-3 Inflammasome Activation and Lipid Accumulation.

J Pharmacol Exp Ther 2016 10 20;359(1):45-53. Epub 2016 Jul 20.

Departments of Drug Science and Technology (E.B., G.V., F.C., M.C., R.F.), Clinical and Biological Sciences (R.M., D.N.), and Biotechnology and Sciences for the Health (J.C.C.), University of Turin, Turin, Italy; and Boehringer Ingelheim Pharma, Biberach an der Riss, Germany (E.M.).

The aim of this study was to evaluate the effects of chronic treatment with empagliflozin, a potent and selective sodium glucose cotransporter-2 inhibitor, in a murine model of diet-induced obesity and insulin resistance, focusing on drug effects on body weight reduction and nucleotide-binding domain, leucine-rich repeat containing protein (NLRP)-3 inflammasome activation, which have never been investigated to date. Male C57BL/6 mice were fed control or a high fat-high sugar (HFHS) diet for 4 months. Over the last 2 months, subsets of animals were treated with empagliflozin (1-10 mg/kg) added to the diet. Empagliflozin evoked body weight reduction (P < 0.001 for the highest dose) and positive effects on fasting glycemia and homeostasis model assessment of insulin resistance. In addition, the drug was able to reduce renal tubular damage and liver triglycerides level in a dose-dependent manner. Interestingly, empagliflozin also decreased cardiac lipid accumulation. Moreover, diet-induced activation of NLRP-3 in kidney and liver (not observed in the heart) was dose-dependently attenuated by empagliflozin. Our results clearly demonstrate the ability of empagliflozin to counteract the deleterious effects evoked by chronic exposure to HFHS diet. Most notably, empagliflozin treatment was associated with NLRP-3 inflammasome signaling modulation, suggesting that this inhibition may contribute to the drug therapeutic effects.
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http://dx.doi.org/10.1124/jpet.116.235069DOI Listing
October 2016

Elevation of serum sphingosine-1-phosphate attenuates impaired cardiac function in experimental sepsis.

Sci Rep 2016 06 9;6:27594. Epub 2016 Jun 9.

The William Harvey Research Institute, Barts &The London School of Medicine &Dentistry, Queen Mary University of London, London, UK.

Serum levels of the lipid mediator sphingosine-1-phosphate (S1P) are reduced in septic patients and are inversely associated with disease severity. We show that serum S1P is reduced in human sepsis and in murine models of sepsis. We then investigated whether pharmacological or genetic approaches that alter serum S1P may attenuate cardiac dysfunction and whether S1P signaling might serve as a novel theragnostic tool in sepsis. Mice were challenged with lipopolysaccharide and peptidoglycan (LPS/PepG). LPS/PepG resulted in an impaired systolic contractility and reduced serum S1P. Administration of the immunomodulator FTY720 increased serum S1P, improved impaired systolic contractility and activated the phosphoinositide 3-kinase (PI3K)-pathway in the heart. Cardioprotective effects of FTY720 were abolished following administration of a S1P receptor 2 (S1P2) antagonist or a PI3K inhibitor. Sphingosine kinase-2 deficient mice had higher endogenous S1P levels and the LPS/PepG-induced impaired systolic contractility was attenuated in comparison with wild-type mice. Cardioprotective effects of FTY720 were confirmed in polymicrobial sepsis. We show here for the first time that the impaired left ventricular systolic contractility in experimental sepsis is attenuated by FTY720. Mechanistically, our results indicate that activation of S1P2 by increased serum S1P and the subsequent activation of the PI3K-Akt survival pathway significantly contributes to the observed cardioprotective effect of FTY720.
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http://dx.doi.org/10.1038/srep27594DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4899780PMC
June 2016