Chloride intracellular channel 1 activity is not required for glioblastoma development but its inhibition dictates glioma stem cell responsivity to novel biguanide derivatives.

BACKGROUND: Chloride intracellular channel-1 (CLIC1) activity controls glioblastoma proliferation. Metformin exerts antitumor effects in glioblastoma stem cells (GSCs) inhibiting CLIC1 activity, but its low potency hampers its translation in clinical settings. METHODS: We synthesized a small library of novel biguanide-based compounds that were tested as antiproliferative agents for GSCs derived from human glioblastomas, in vitro using 2D and 3D cultures and in vivo in the zebrafish model. Compounds were compared to metformin for both potency and efficacy in the inhibition of GSC proliferation in vitro (MTT, Trypan blue exclusion assays, and EdU labeling) and in vivo (zebrafish model), migration (Boyden chamber assay), invasiveness (Matrigel invasion assay), self-renewal (spherogenesis assay), and CLIC1 activity (electrophysiology recordings), as well as for the absence of off-target toxicity (effects on normal stem cells and toxicity for zebrafish and chick embryos). RESULTS: We identified Q48 and Q54 as two novel CLIC1 blockers, characterized by higher antiproliferative potency than metformin in vitro, in both GSC 2D cultures and 3D spheroids. Q48 and Q54 also impaired GSC self-renewal, migration and invasion, and displayed low systemic in vivo toxicity. Q54 reduced in vivo proliferation of GSCs xenotransplanted in zebrafish hindbrain. Target specificity was confirmed by recombinant CLIC1 binding experiments using microscale thermophoresis approach. Finally, we characterized GSCs from GBMs spontaneously expressing low CLIC1 protein, demonstrating their ability to grow in vivo and to retain stem-like phenotype and functional features in vitro. In these GSCs, Q48 and Q54 displayed reduced potency and efficacy as antiproliferative agents as compared to high CLIC1-expressing tumors. However, in 3D cultures, metformin and Q48 (but not Q54) inhibited proliferation, which was dependent on the inhibition dihydrofolate reductase activity. CONCLUSIONS: These data highlight that, while CLIC1 is dispensable for the development of a subset of glioblastomas, it acts as a booster of proliferation in the majority of these tumors and its functional expression is required for biguanide antitumor class-effects. In particular, the biguanide-based derivatives Q48 and Q54, represent the leads to develop novel compounds endowed with better pharmacological profiles than metformin, to act as CLIC1-blockers for the treatment of CLIC1-expressing glioblastomas, in a precision medicine approach.

aEEG in preterm infants.

Over the last decade, the survival of premature babies has improved dramatically. Such infants, especially those with extremely low birth weight, are still affected by dangerous complications occurring during the neonatal period that often cause brain damage. Intraventricular-intraparenchymal haemorrhage (IVH-IPH), periventricular leukomalacia (PVL), seizures, meningitis and hypoxic-ischaemic encephalopathy are the most common complications. Such problems require more specialized monitoring of brain function during this critical period. In recent years, many studies on very premature infants have shown that aEEG has a high predictive value for both short-term and long-term outcome. In fact, it has been proven that some types of background activity patterns, the absence of a sleep-wake cycle, and seizure activity are related to the onset of early complications such as IVH-IPH and PVL. Most recent studies have shown that an aEEG performed in the early hours or during the first days of life can predict the neurobehavioural development of preterm infants at 2 years and 3 years (Bayley Scale). In particular our study demonstrates that loss of sleep-wake cycling, shown by aEEG, has a high positive predictive value for the development of posthaemorrhagic hydrocephalus (PPH) in preterm infants with IVH; therefore, the study of cerebral background activity and in particular of sleep-wake cycling can be used as an early prognostic tool in patients at risk of PPH.

The role of probiotics in the clinical management of food allergy and atopic dermatitis.

The probiotics are cultures of potentially beneficial bacteria that positively affects host by enhancing the microbial balance and therefore restore the normal intestinal permeability and gut microecology, improve the intestine's immunologic barrier function and reduce the generation of proinflammatory cycochines characteristics of allergic inflammation. The authors made a survey of the most relevant studies concerning the use of probiotics in the food allergy, atopic dermatitis, and in primary prevention of atopy. On the basis of these trials probiotic therapy alleviates allergic inflammation as demonstrated by the control of clinical symptoms and the reduction of local and systemic inflammatory markers.

Probiotics and atopic dermatitis. A new strategy in atopic dermatitis.

Over the last few decades, the prevalence of atopic dermatitis has been increasing from 2% to 100%, with 90% of cases within 5 years of age versus 6% between 6 and 10 years and 2% after 10 years, and environmental factors may possibly play an important role in this increase as in other atopic diseases. Many findings suggest an important role of atopy in atopic dermatitis; moreover, 40% of children with atopic dermatitis have food allergy and the removal of the food allergen from the patient's diet leads to a significant clinical improvement. In a possible scenario, IgE-bearing dendritic cells are likely to process allergens acquired in the gastrointestinal tract, circulate to the skin and activate local T cells. Cultures of beneficial live microorganisms characteristic of the commensal microflora are administered with probiotic functional foods in order to provide a microbial challenge for the maturation of gut-associated lymphoid tissue, which the infant often lacks. The probiotic effects are attributed to normalisation of the increased intestinal permeability and balancing gut microecology, improvement of the immunological defence barrier (IgA) of the intestine, alleviation of the intestinal inflammatory response, and downregulation of proinflammatory cytokines characteristic of local and systemic allergic inflammation.

Esophageal hyperalgesia in patients with ulcerative colitis: role of experimental stress.

OBJECTIVES: Intestinal inflammation is associated with enteric nervous system alterations, at both inflamed and noninflamed sites. The perception of stimuli from the GI tract is enhanced during inflammatory conditions, but it is unknown whether visceral hypersensitivity is limited to the inflamed area or diffuse throughout the entire GI tract. Moreover, although stress can reactivate inflammatory processes in the gut, it is unknown if this can alter perception from the GI tract. Our aim was to determine if patients with ulcerative colitis (UC) have increased esophageal sensitivity to distention and whether this is modified by experimental stress. METHODS: Ten UC patients and 12 healthy volunteers (HVs) underwent gradual balloon distension of the esophagus to assess their visceral sensitivity. Perceptive and pain thresholds were evaluated in basal conditions and after induction of experimental stress (cold water pressure test) while blood pressure and heart rate were monitored. RESULTS: Patients with UC had perceptive thresholds to distension similar to HVs (14.8+/-2.0 ml of air vs 14.5+/-3.0 ml); in contrast, the volume increment needed to evoke pain was significantly lower in UC patients than in HVs (58.9% vs 149.9%, p < 0.05). Physical stress caused a similar decrease in perceptive thresholds in HVs (-29.1+/-8.4%) and patients (-17.7+/-9.1%), but pain thresholds were significantly decreased only in HVs (-28.3+/-7.1% vs -11.5+/-12.3%). CONCLUSIONS: UC is characterized by increased esophageal sensitivity, indicating the existence of diffuse hyperalgesia during intestinal inflammatory processes. This increased sensitivity may account for the frequent upper GI symptoms these patients complain of when in clinical remission.

Long term efficacy, safety, and tolerabilitity of low daily doses of isosmotic polyethylene glycol electrolyte balanced solution (PMF-100) in the treatment of functional chronic constipation.

AIMS: To assess the long term therapeutic effectiveness, safety, and tolerability of low daily doses of isosmotic PEG electrolyte solutions (PMF-100) administered for a six month period for the treatment of functional constipation, in a double blind, placebo controlled, parallel group study. METHODS: After an initial four week run in period with PMF-100 (250 ml twice daily; PEG 14.6 g twice daily), 70 patients suffering from chronic constipation (58 females, aged 42 (15) years) with normalised bowel frequency (>3 bowel movements (bm)/week) were randomly allocated to receive either PMF-100 or placebo, contained in sachets (one sachet in 250 ml of water twice daily) for 20 weeks. Patients were assessed at four week intervals, and reported frequency and modality of evacuation, laxative use, and relevant symptoms on a diary card. At weeks 1, 12, and 24, a physical examination and laboratory tests were performed. RESULTS: Complete remission of constipation was reported by a significantly (p<0.01) higher number of patients treated with PMF-100 compared with placebo at each four week visit. At the end of the study, 77% of the PMF-100 group and 20% of the placebo group were asymptomatic. Compared with placebo, patients treated with PMF-100 reported hard/pellety stools and straining at defecation less frequently, a significantly higher bowel frequency (week 12: 7. 4 (3.1) v 4.3 (2.5) bm/week, 95% CI 1.64, 4.42; week 24: 7.4 (3.2) v 5.4 (2.1) bm/week, 95% CI 0.13,3.93), reduced consumption of laxative/four weeks (week 12: 0.7 (2.7) v 2.2 (3.3), 95% CI -2.29, 0. 03; week 24: 0.2 (0.8) v 1.4 (2), 95% CI -2.07, -0.023), reduced mean number of sachets used (week 12: 33 (13) v 43 (12), 95% CI -17. 24, 4.56; week 24: 33 (13) v 44 (12), 95% CI -19.68, -2.24), and reduced number of drop outs for therapy failure (16 v 3; p<0.005). Adverse events, physical findings, laboratory values, palatability, and overall tolerance of the solutions did not differ between groups. CONCLUSIONS: Administration of small daily doses of isosmotic PEG electrolyte balanced solutions was effective over a six month period for the treatment of functional constipation. A mean daily dose of approximately 300 ml of PEG solution (PEG 17.52 g) appeared to be safe, well tolerated, and devoid of significant side effects.

[Collection, survey and interpretation of pH-metry data]. / Rilievo, acquisizione ed elaborazione del dato pH-metrico.

24-hour oesophageal pH monitoring is a necessary tool to diagnose Gastro-Oesophageal Reflux Disease. The technical characteristics of the available equipments have been improved and standard criteria to identify reflux episodes have been defined, increasing the sensitivity and specificity of the test. The state of the art on the technical features of prolonged pHmetry will be reviewed in this paper.