Circulating miRNA 122-5p Expression Predicts Mortality and Cardiovascular Events in Chronic Hemodialysis Patients: A Multicentric, Pilot, Prospective Study.

BACKGROUND: Despite patients undergoing chronic hemodialysis (HD) being notoriously prone to adverse cardiovascular (CV) events, risk prediction in this population remains challenging. miRNA 122-5p, a short, non-coding RNA predominantly involved in lipid and carbohydrate metabolism, has recently been related to the onset and progression of CV disease. METHODS: We run a pilot, multicenter, longitudinal, observational study to evaluate the clinical significance and prognostic usefulness of circulating miRNA 122-5p in a multicentric cohort of 74 individuals on maintenance HD. RESULTS: Patients displayed lower circulating miRNA 122-5p as compared to healthy controls (p = 0.004). At correlation analyses, ALT (ß = 0.333; p = 0.02), E/e' (ß = 0.265; p = 0.02) and CRP (ß = -0.219; p = 0.041) were independent predictors of miRNA 122-5p levels. During a median follow-up of 22 months (range of 1-24), 30 subjects (40.5%) experienced a composite endpoint of all-cause mortality and fatal/non-fatal CV events. Baseline circulating miRNA 122-5p was higher in these subjects (p = 0.01) and it predicted a significantly higher risk of endpoint occurrence (Kaplan-Meier crude HR 3.192; 95% CI 1.529-6.663; p = 0.002; Cox regression adjusted HR 1.115; 95% CI 1.009-1.232; p = 0.03). CONCLUSIONS: Altered miRNA 122-5p levels in HD patients may reflect hepatic and CV damage and may impart important prognostic information for improving CV risk prediction in this particular setting.

Amyloid-ß-Induced Transglutaminase 2 Expression and Activities are Modulated by 2-Pentadecyl-2-Oxazoline in Mouse and Human Microglial Cell Lines.

BACKGROUND: Transglutaminase 2 is an ubiquitously multifunctional enzyme and the most widely studied of the transglutaminase family. Consistent with its role in promoting post-translational modifications of proteins, Transglutaminase 2 is involved in many physiological processes such as apoptosis, signal transduction, and cellular adhesion. Several findings indicate that Transglutaminase 2 plays a role in the pathological processes of various inflammation-related diseases, including neurodegenerative diseases. OBJECTIVE: We tested the potential modulatory effects on amyloid-ß-induced Transglutaminase 2 expression and activities of 2-pentadecyl-2-oxazoline, a plant-derived agent, which has shown effectiveness against chronic pain and associated neuropsychiatric disorders, both in mouse and human microglial cell lines. METHODS: We used biochemistry, molecular and cell biology techniques to evaluate the potential modulatory effects on amyloid-ß-induced Transglutaminase 2 expression and activities of 2- pentadecyl-2-oxazoline in mouse and human microglial cell lines. RESULTS: 2-pentadecyl-2-oxazoline was able to modulate amyloid-ß-induced Transglutaminase 2 expression and activities in mouse and human microglial cell lines. CONCLUSION: Transglutaminase 2 confirms its role as a neuroinflammation marker, the inhibition of which could be a potential preventive and therapeutic approach, while 2-pentadecyl-2-oxazoline is a potent modulator of the amyloid-ß-induced Transglutaminase 2 expression and activities in mouse and human microglial cell lines.

Brain Growth Evaluation Assessed with Transfontanellar (B-GREAT) Ultrasound. Old and New Bedside Markers to Estimate Cerebral Growth in Preterm Infants: a Pilot Study.

OBJECTIVE: We aimed to investigate the feasibility of evaluating overall preterm brain growth using a gathered set of measurements of brain structures in standard cranial ultrasound planes. We called this method of assessment Brain Growth Evaluation Assessed with Transfontanellar ultrasound (B-GREAT). STUDY DESIGN: In this prospective observational cohort study, cranial ultrasound was regularly performed (on day 1, 2, 3, and 7 of life, and then weekly until discharge, and at term) in preterm infants born with gestational age (GA) less than 32 weeks. We evaluated corpus callosum length, corpus callosum-fastigium length, anterior horn width, frontal white matter height, total brain surface, deep grey matter height, hemisphere height, transverse cerebellar diameter in the axial view, and transverse cerebellar diameter coronal view. Measurements obtained were used to develop growth charts for B-GREAT markers as a function of postmenstrual age. Reproducibility of B-GREAT markers was studied. RESULTS: A total of 528 cranial ultrasounds were performed in 80 neonates (median birth GA: 28+5 weeks and interquartile range: 27+3-30+5). The intraclass correlation coefficients for intra-observer and inter-observer analyses showed substantial agreement for all B-GREAT markers. Growth curves for B-GREAT markers were developed. CONCLUSION: B-GREAT is a feasible and reproducible method for bedside monitoring of the growth of the main brain structures in preterm neonates. KEY POINTS: · Overall neonatal brain growth is not routinely monitored using ultrasound.. · Old and new markers were used to build a standardized and non-invasive tool to monitor brain growth.. · All B-GREAT measurements had a good intra-observer and inter-observer agreement..

Altered circulating marinobufagenin levels and recurrent intradialytic hypotensive episodes in chronic hemodialysis patients: a pilot, prospective study.

Intradialytic hypotension (IDH) is a sudden and often serious complication of chronic hemodialysis (HD). In this prospective study, we aimed at evaluating the clinical predictors of IDH in a homogeneous cohort of chronic HD patients, with a particular focus on marinobufagenin (MBG), an endogenous cardiotonic steroid which alterations have previously been involved in various cardiovascular disorders. MBG levels in HD patients were significantly higher than in controls (p = 0.03), remained unchanged throughout a single HD session and were not correlated with the absolute or partial fluid loss achieved. During a 30-day follow-up, 19 patients (65.5%) experienced at least one IDH (73 total episodes). An inverse correlation was found between baseline MBG and the number of IDH (R = -0.55; p = 0.001). HD patients experiencing IDH presented remarkably lower baseline MBG as compared to others (p = 0.008) with a statistically significant trend during HD (p = 0.02). At Kaplan-Meier analyses, HD patients with lower MBG manifested a four-to-six fold increased risk of IDH during follow-up (crude Hazard Ratio ranging from 4.37 to 6.68). At Cox regression analyses, MBG measurement at different time points resulted the strongest time-dependent predictors of IDH among all the variables considered (HR ranging from 0.068 to 0.155; p: 0.002 to <0.0001). Findings obtained suggest that differently altered MBG in chronic HD patients may reflect a diverse vascular and hemodynamic tolerance to HD stress, eventually leading to recurrent IDH episodes. Further studies are needed to confirm the prognostic capacity of MBG for identifying HD patients at high risk of IDH, particularly those with apparently optimal fluid status.

Perturbations of the hepatic proteome behind the onset of metabolic disorders in mouse offspring developed following embryo manipulation.

Assisted Reproductive Technologies (ART) allowed the births of >8 million babies worldwide. Even if ART children are healthy at birth, several studies reported that ART may cause changes in foetal programming, leading to an increased predisposition to metabolic disorders in adulthood. Previous studies on mouse model showed obesity, glucose intolerance, and hepatic lipid accumulation in ART offspring. A cumulative effect of the different components of ART protocol has been previously described, for example, in the occurrence of epigenetic defects. Here, we investigated whether there is a cumulative effect of embryo transfer (ET), in vitro culture (IVC) and blastomere biopsy (BB) in the onset of metabolic disorders in mouse offspring vs those naturally conceived (Control - CTR). To this aim, proteomic analysis was performed on the livers from adult mouse offspring developed following ET, IVC and BB vs CTR. We observed deregulated expression of proteins involved in lipid, carbohydrate, energy metabolisms and cellular processes in ART offspring. Moreover, we found increased body weight in all ART offspring while i) insulin resistance in BB male, ii) females glucose intolerance and high level of triglycerides and cholesterol in BB females and iii) low levels of interleukin-6 in BB, IVC and ET males. In conclusion, our study suggests that the use of various embryo manipulations influences the metabolic health of adult offspring, resulting in an increased predisposition to hepatic diseases and metabolic syndrome in a sex-specific manner.

Lipid droplets in mammalian eggs are utilized during embryonic diapause.

Embryonic diapause (ED) is a temporary arrest of an embryo at the blastocyst stage when it waits for the uterine receptivity signal to implant. ED used by over 100 species may also occur in normally "nondiapausing" mammals when the uterine receptivity signal is blocked or delayed. A large number of lipid droplets (LDs) are stored throughout the preimplantation embryo development, but the amount of lipids varies greatly across different mammalian species. Yet, the role of LDs in the mammalian egg and embryo remains unknown. Here, using a mouse model, we provide evidence that LDs play a crucial role in maintaining ED. By mechanical removal of LDs from zygotes, we demonstrated that delipidated embryos are unable to survive during ED. LDs are not essential for normal prompt implantation, without ED. We further demonstrated that with the progression of ED, the amount of intracellular lipid reduces, and composition changes. This decrease in lipid is caused by a switch from carbohydrate metabolism to lipid catabolism in diapausing blastocysts, which also exhibit increased release of exosomes reflecting elevated embryonic signaling to the mother. We have also shown that presence of LDs in the oocytes of various mammals positively corelates with their species-specific length of diapause. Our results reveal the functional role of LDs in embryonic development. These results can help to develop diagnostic techniques and treatment of recurrent implantation failure and will likely ignite further studies in developmental biology and reproductive medicine fields.

Blastomere removal affects homeostatic control leading to obesity in male mice.

Preimplantation embryos are particularly vulnerable to environmental perturbations, including those related to assisted reproductive technologies. Invasive embryo manipulations, such as blastomere biopsy, are applied worldwide in clinical settings for preimplantation genetic testing. Mouse models have previously shown that blastomere biopsy may be associated with altered phenotypes in adult offspring. The aim of the present study was to investigate the specific contribution of blastomere removal to the physiological, behavioral, and molecular regulators of energy homeostasis, as compared to sham manipulation (re-introducing the blastomere into the embryo after its removal) and in vitro culture. Mice derived from 8-cell embryos subjected to blastomere removal displayed: (i) higher body weight and adiposity, (ii) increased food intake and sucrose preference, (iii) decreased time of immobility in the tail suspension test, and (iv) resistance to weight loss after social isolation or following 3 days of physical exercise - compared to mice derived from sham biopsy or from in vitro-cultured embryos. Mice generated after blastomere removal also had increased circulating leptin and leptin gene expression in adipose tissue, as well as increased ghrelin receptor gene expression in the hypothalamus, compared to control mice. The effects of blastomere biopsy on offspring phenotype were sexually dimorphic, with females not being affected. These results indicate that blastomere deprivation, rather than other perturbations of the blastomere biopsy procedure, programs male embryos to develop physiological, behavioral, and molecular dysregulation of energy homeostasis, leading to postnatal obesity.

Neurodevelopmental outcomes in very preterm infants: The role of severity of Bronchopulmonary Dysplasia.

OBJECTIVE: Bronchopulmonary dysplasia is a chronic respiratory disease that still affects preterm neonates; its association with neurodevelopmental (ND) impairment is already known. Different studies investigated neurodevelopmental outcomes in infants with BPD, often using the old dichotomous definition (BPD vs Non-BPD). This retrospective study aims to evaluate the role of different BPD severity grades on ND outcomes at 24 months of corrected age (CA). METHODS: All preterm infants born between 2011 and 2015 in the study hospital with a gestational age (GA) ≤ 30 weeks and discharged from our NICU were included and were divided in infants with and without BPD. Infants with BPD were divided into three severity groups as defined by NICHD/NHLBI Workshop in 2001, and were compared to their Non-BPD peers, matching them according to the same GA and year of birth. At 24 months postmenstrual age, we assessed general outcomes (growth and hospital readmissions) and neurodevelopmental outcomes (motor, developmental and sensory outcomes) with a standardized assessment. RESULTS: We enrolled 89 patients affected by BPD of different grades of severity and a control group of 89 preterm infants without BPD. Infants with Moderate and Severe BPD showed a significantly higher corrected odds ratio (OR) for cognitive impairment compared to controls. Within the group of infants without severe disability (regarding Griffiths' scales), infants with Moderate and Severe BPD as well as infants with Mild BPD showed a significantly higher risk of a lower total Developmental Quotient (DQ) score, even after correction for confounding factors. CONCLUSIONS: Our study evidenced that not only Severe BPD infants, but also Moderate ones showed a higher risk of overall cognitive impairment at 24 months CA. Within the group of infants without severe disability, also those with Mild BPD had lower Griffiths DQ scores than those without. This would suggest that infants with BPD, regardless of severity, warrant neurodevelopmental follow-up.

Mouse single oocyte imaging by MALDI-TOF MS for lipidomics.

Reproductive cells are a very special kind of material for the analysis. Depending on the species, their dimensions allow for the application of mass spectrometry imaging-based techniques to receive a reasonable data for interpretation of their condition without any additional sample preparation steps, except for typical sample preparation characteristic for IMS protocols. A comparison between lipid profiles of oocytes could answer the question of the overall quality of the cells in the function of time or conditions of storage. Even tiny differences in the lipid profiles, but still detectable by bioinformatic analysis, could be crucial for the estimation of the conditions of the cells in various stages of development or aging. In our study, MALDI-TOF/TOF MSI was used to analyze and visualize the single oocytes. We deposited the cells on the transparent indium-tin-oxide (ITO) glass and marked their positions, which allowed for the fast localization of the cells and precise laser targeting in the ion source. We also optimized the usage of different MALDI matrices and different approaches. The proposed way of measurement allows analyzing quite a significant quantity of oocytes in a reasonably short time. During the analysis, the lipid composition of the single cell was successfully estimated in a conventional usage of the MALDI ion source, and the localization of lipids was confirmed by imaging mass spectrometry (IMS) analysis. The observed quantity of the lipids allowed for the application of the LIFT™ technique to obtain MS/MS spectra sufficient for lipids' unambiguous identification. We hope that our idea of the oocyte analysis will help to elucidate chemical changes that accompany different processes in which oocytes are involved. There could be such fascinating phenomena as the oocyte maturation, changes in the lipid components during their storage, and much more.

Lung ultrasonography score versus chest X-ray score to predict surfactant administration in newborns with respiratory distress syndrome.

OBJECTIVES: We aim to verify the diagnostic accuracy of a lung ultrasonography (LUS) score to early predict the need for surfactant therapy in newborns with respiratory distress syndrome (RDS), and to compare it with a chest X-ray score. METHODS: In this prospective diagnostic accuracy study we included all newborns admitted for respiratory distress and initially treated with nasal CPAP. LUS was performed within 2 h from nasal CPAP positioning and in any case before surfactant administration. A chest X-ray was also performed. A LUS score and an X-ray score were used and compared. Ability of the scores to predict surfactant administration was evaluated through ROC analysis. RESULTS: In our population of 56 newborns with mean gestational age of 31 weeks (SD 3) and mean birth weight of 1442 g (SD 520), LUS score showed higher AUC than X-ray score in early recognition of infants with respiratory distress syndrome requiring surfactant treatment (0.94; 95%CI, 0.89-0.98; P < 0.001 vs 0.80; 95%CI, 0.74-0.86; P < 0.001). It showed also higher sensitivity (86% vs 82%), higher specificity (88% vs 76%), better positive (83% vs 69%), and negative (91% vs 87%) predictive values. CONCLUSIONS: LUS is a non-invasive, bedside and reproducible method that could improve the management of neonatal respiratory distress. It is accurate and reliable to early identify patients who will need treatment with surfactant allowing both an early treatment and a reduction of radiation exposure.

Developmental peculiarities in placentae of ovine uniparental conceptuses.

Genomic imprinting is an epigenetic phenomenon regulating mono-allelic expression of genes depending on their parental origin. Defective genomic imprinting is involved in several placental disorders, such as intrauterine growth restriction and pre-eclampsia. Uniparental embryos, having maternal-only or paternal-only genomes (parthenogenotes [PAR] and androgenotes [AND], respectively), are useful models to study placentation. The aim of this work was to reveal the effect of parental genome (maternal and paternal) on placentation. To do this, uniparental (AND and PAR) and biparental (CTR) in vitro produced sheep embryos transferred to recipient females were collected at day 20 of pregnancy and their placentae were analyzed. qPCR analysis showed that imprinted genes (H19, IGF2R and DLK1) were expressed accordingly to their parental origin while the expression f DNA methyltransferases () was disregulated, especially in PAR (P < 0.05). AND placentae were significantly hypomethylated compared to both PAR and CTR (P = 0.023). Chorion-allantoid of AND showed impaired development of vessels and reduced mRNA expression of vasculogenetic factors (ANG2 P = 0.05; VEGFR2 P< 0.001; TIE2 P < 0.001). Morphologically, PAR placentae were characterized by abnormal structure of the trophoectodermal epithelium and reduced total number (P<0.03) of Trophoblastic Binucleate Cells. A reduced implantation rate of both classes of uniparental embryos (P<0.03) was also noted. Our results provide new insights into the characterization of uniparental embryos and demonstrate the complementary role of parental genomes for the correct establishment of pregnancy. Thus, our findings may suggest new targets to improve our understanding of the origin of imprinting-related placental dysfunction.

Embryo biopsy and development: the known and the unknown.

Preimplantation genetic diagnosis (PGD) has been introduced in clinical practice as a tool for selecting 'healthy' embryos before their transfer in utero. PGD protocols include biopsy of cleaving embryos (blastomere biopsy (BB)) or blastocysts (trophectoderm biopsy (TB)), followed by genetic analysis to select 'healthy' embryos for transfer in utero. Currently, TB is replacing the use of BB in the clinical practice. However, based on the European Society of Human Reproduction and Embryology Preimplantation Genetic Diagnosis Consortium reports, BB has been used in >87% of PGD cycles for more than 10 years. An exhaustive evaluation of embryo biopsy (both BB and TB) risks and safety is still missing. The few epidemiological studies available are quite controversial and/or are limited to normalcy at birth or early childhood. On the other hand, studies on animals have shown that BB can be a risk factor for impaired development, during both pre- and postnatal life, while little is known on TB. Thus, there is an urgent need of focused researches on BB, as it has contributed to give birth to children for more than 10 years, and on TB, as its application is significantly growing in clinical practice. In this context, the aim of this review is to provide a complete overview of the current knowledge on the short-, medium- and long-term effects of embryo biopsy in the mouse model.

Constipation and cow's milk allergy: a review of the literature.

The causal association between cow's milk allergy (CMA) and constipation is not well established. Some guidelines describe constipation as a possible symptom of CMA, while others do not mention it. We conducted a literature review and found 10 prospective clinical trials. In all of them, an oral food challenge was performed, and 2 of them were randomized. These studies reported that a cow's milk (CM) protein-free diet has a beneficial effect on constipation, with a rate of successful outcomes ranging from 28 to 78%. The hypothetic pathogenic mechanism lies in increased anal pressure at rest, probably caused by allergic inflammation of the internal sphincter area due to mucosal eosinophil and mast cell infiltration. Eighty percent of patients reach tolerance within 1 year after the diagnosis of CMA-related constipation. We believe that a CM-free diet for 2-4 weeks should be proposed for children with chronic functional constipation, even if it is not severe or resistant to laxatives.

Platinum compounds in children with cancer: toxicity and clinical management.

Platinum compounds are widely used in the treatment of pediatric tumors such as neuroblastoma, germ-cell tumors, osteosarcoma, retinoblastoma, hepatoblastoma, brain tumors (low-grade gliomas and medulloblastoma/PNET), and relapsed and refractory lymphomas. The three major platinum compounds (cisplatin, carboplatin, and oxaliplatin) have a similar pharmacokinetics profile and mechanism of action, but the differences in their chemical structure are responsible for their different antitumor activity and toxicity. In this review, we have described the main characteristics of cisplatin, carboplatin, and oxaliplatin, focusing on their toxic effects and possible strategies to prevent them to improve the clinical outcomes in pediatric cancer patients. The underlying mechanism of each platinum-related toxicity is shown together with the clinical manifestations. Furthermore, possible preventive strategies are suggested to reduce the negative impact of platinum compounds on the quality of life of children with cancer. Cisplatin seems to be mostly ototoxic and nephrotoxic, carboplatin mainly produces myelosuppression, whereas oxaliplatin induces predominantly peripheral sensory neurotoxicity. In contrast, nausea and vomiting can be linked to all platinum compounds, although cisplatin exerts the strongest emetic effect. A correct knowledge of pharmacokinetics and toxicological profile of platinum compounds may aid physicians prevent their toxicity on auditory, nervous, renal, and bone marrow function, improving the quality of life of pediatric cancer patients.

Azole interactions with multidrug therapy in pediatric oncology.

Patients with cancer receive multidrug therapy. Antineoplastic agents and supportive care drugs are often administered together, leading to potential drug-drug interactions. These interactions may have significant clinical implications in terms of toxicity or a decrease in the efficacy of the treatment administered. Here, we focus on the role of azoles and their main pharmacokinetic interactions with the principal classes of drugs used in pediatric oncology. The co-administration of azoles and antineoplastic agents, corticosteroids, immunosuppressants, antacids, antiemetics, antiepileptic drugs and analgesics was investigated, and a practical guide on the management of these drugs when administered together is provided.

Efficacy and safety of transdermal buprenorphine in the management of children with cancer-related pain.

BACKGROUND: The current study investigated the efficacy, safety, tolerability, and compliance of a transdermal buprenorphine delivery system for the management of chronic cancer pain in the pediatric population. PROCEDURE: Sixteen pediatric patients with moderate to severe cancer-related pain not satisfactorily controlled with previous non-opioid therapies were enrolled. Transdermal buprenorphine was administered following a 72 hour schedule and rescue medication (tramadol) was allowed for breakthrough pain. Pain intensity was assessed using the Wong-Baker faces pain rating scale (WBS) and other parameters related to the global quality of life were evaluated. Children's evaluations of efficacy, compliance, and tolerability were recorded using numerical scales. Adverse events were monitored during the study and the medications needed to control opioid-related nausea and constipation were recorded. RESULTS: Eleven patients (68.75%) responded to transdermal buprenorphine after 2 weeks of treatment. Pain intensity measured with WBS decreased from 6.25 at baseline to 1.38 at Day +60 (P < 0.001). All outcome measures of global quality of life (quality of sleep, alimentation, play and activity, speech, and crying) significantly improved over the 60-day study period. Children's evaluations of compliance and tolerability of the drug were always positive over the entire period of treatment. No severe adverse events were recorded. Opioid-related nausea was well controlled with medication on request, and the need for laxative therapy was greater at the end of the second month of treatment. CONCLUSIONS: Transdermal buprenorphine was found to represent an efficient, safe and well tolerated approach to the management of children's chronic cancer pain.