Regulation of early steps of GPVI signal transduction by phosphatases: a systems biology approach

We present a data-driven mathematical model of a key initiating step in platelet activation, a central process in the prevention of bleeding following Injury. In vascular disease, this process is activated inappropriately and causes thrombosis, heart attacks and stroke. The collagen receptor GPVI is the primary trigger for platelet activation at sites of injury. Understanding the complex molecular mechanisms initiated by this receptor is important for development of more effective antithrombotic medicines. In this work we developed a series of nonlinear ordinary differential equation models that are direct representations of biological hypotheses surrounding the initial steps in GPVI-stimulated signal transduction. At each stage model simulations were compared to our own quantitative, high-temporal experimental data that guides further experimental design, data collection and model refinement. Much is known about the linear forward reactions within platelet signalling pathways but knowledge of the roles of putative reverse reactions are poorly understood. An initial model, that includes a simple constitutively active phosphatase, was unable to explain experimental data. Model revisions, incorporating a complex pathway of interactions (and specifically the phosphatase TULA-2), provided a good description of the experimental data both based on observations of phosphorylation in samples from one donor and in those of a wider population. Our model was used to investigate the levels of proteins involved in regulating the pathway and the effect of low GPVI levels that have been associated with disease. Results indicate a clear separation in healthy and GPVI deficient states in respect of the signalling cascade dynamics associated with Syk tyrosine phosphorylation and activation. Our approach reveals the central importance of this negative feedback pathway that results in the temporal regulation of a specific class of protein tyrosine phosphatases in controlling the rate, and therefore extent, of GPVI-stimulated platelet activation

Evidence for the presence of non-celiac gluten sensitivity in patients with functional gastrointestinal symptoms : Results from a multicenter randomized double-blind placebo-controlled gluten challenge

Non-celiac gluten sensitivity (NCGS) is characterized by the onset of symptoms after eating gluten-containing food. We aimed to single out NCGS subjects among subjects with functional gastrointestinal symptoms. Patients were enrolled in a multicenter double-blind placebo-controlled trial with crossover. Symptoms and quality of life were evaluated by means of 10-cm VAS and SF36. Iron parameters, transaminases and C reactive protein (CRP) were evaluated. After a three-week-long gluten-free diet (GFD), responsive patients were randomly assigned to gluten intake (5.6 g/day) or placebo for seven days, followed by crossover. The primary endpoint was the worsening of symptoms (VAS increase 653 cm) during gluten ingestion compared to placebo. One hundred and forty patients were enrolled and 134 (17 males, mean age 39.1 \ub1 11.7 years, BMI 22.4 \ub1 3.8) completed the first period. A total of 101 subjects (10 males, mean age 39.3 \ub1 11.0 years, BMI 22.3 \ub1 4.0) reported a symptomatic improvement (VAS score 2.3 \ub1 1.2 vs. 6.5 \ub1 2.2 before and after GFD, p = 0.001). 98 patients underwent the gluten challenge and 28 (all females, mean age 38.9 \ub1 12.7 years, BMI 22.0 \ub1 2.9) reported a symptomatic relapse and deterioration of quality of life. No parameters were found to be statistically associated with positivity to the challenge. However, 14 patients responded to the placebo ingestion. Taking into account this finding, about 14% of patients responding to gluten withdrawal showed a symptomatic relapse during the gluten challenge. This group is suspected to have NCGS

Molecular characterization of Canine minute virus associated with neonatal mortality in a litter of Jack Russell terrier dogs

The molecular characterization of a strain of Canine minute virus (CnMV) associated with neonatal death is reported. Three newborn puppies of a litter of Jack Russell terrier dogs died after displaying systemic disease, whereas 2 surviving puppies showed no clinical signs with the exception of transient cardiac abnormalities that were evident by electrocardiography. Necropsy of 1 dead puppy revealed severe lesions in the internal organs. A strain of Canine minute virus was detected in tissue samples collected from the puppy, and virus circulation was demonstrated by molecular or serological testing in the dam, puppies of the same litter, and other puppies in the same kennel. By sequence and phylogenetic analysis of the gene encoding for the VP2 capsid protein, the strain circulating in the kennel was found to be related to recent Asian CnMV isolates. Continuous molecular surveillance for CnMV in kennels, shelters, and rescue centers would expand the knowledge base on the epidemiological and pathogenetic features of CnMV, which has been known for several decades but still poorly understoo

Base genetica della microangiopatia trombotica post-trapianto di midollo: l’ultimo tassello del puzzle? [The genetic basis of post-bone marrow transplant thrombotic microangiopathy: is this the last piece of the puzzle?]

La microangiopatia trombotica associata al trapianto (TA-TMA) è una complicanza del trapianto di cellule staminali ematopoietiche (HSCT) correlata a danno renale e significativa mortalità. Studi recenti indicano che la disregolazione della via alterna del complemento potrebbe essere alla base dello sviluppo di TATMA. Attualmente, non esistono strumenti di screening pre-trapianto per identificare i pazienti a rischio. Riportiamo il caso clinico di una donna di 47 anni, sottoposta a HSCT aploidentico complicato da TA-TMA confermato dalla biopsia renale. Per esplorare il meccanismo della TA-TMA, abbiamo eseguito uno studio genetico che ha permesso di identificare la delezione della regione CFHR3-CFHR1 in omozigosi. La paziente sospendeva il trattamento con inibitori della calcineurina (potenziali induttori di TA-TMA) con temporanea introduzione di prednisone fino a completa risoluzione del danno renale e della microangiopatia. L’identificazione di varianti genetiche che interessano il meccanismo della via alterna del complemento potrebbe essere d’ausilio nella stratificazione del rischio di TA-TMA e nella personalizzazione dell’approccio terapeutico.Transplant-associated thrombotic microangiopathy (TA-TMA) is a complication of hematopoietic stem cell transplantation (HSCT) associated with kidney injury and significant mortality. Recent studies indicate that dysregulation of the alternate complement pathway may be at the basis of the development of TA-TMA. Currently, there are no pre-transplant screening tools to identify patients at risk. To explore the mechanism of TA-TMA, we performed a genetic study that allowed us to identify the deletion of the CFHR3-CFHR1 region in homozygosity. We report the clinical case of a 47-year-old woman who underwent haploidentical HSCT complicated by TA-TMA confirmed by renal biopsy. The patient discontinued treatment with calcineurin inhibitors (potential inducers of TA-TMA) with a brief introduction of prednisone until complete resolution of renal damage and microangiopathy. Identifying genetic variants that affect the mechanism of the alternate complement pathway could help in the stratification of the risk of TA-TMA and in implementing a personalized therapeutic approach. Copyright by Società Italiana di Nefrologia SIN, Rome, Italy

Hyoscine N-butylbromide for adenoma detection during colonoscopy : a randomized, double-blind, placebo-controlled study

Background: Hyoscine N-butylbromide (HBB), commonly used during colonoscopy to facilitate cecal intubation, has been proposed to increase the adenoma detection rate (ADR). Aims: To evaluate whether HBB administration increases the adenoma detection rate and influences patients' tolerance. Methods: Consecutive colonoscopy outpatients were randomized after cecal intubation to receive either 20. mg HBB or placebo i.v. The number, size, histology and location of polyps were recorded. The air retained in the abdomen was either indirectly estimated by \u3b4AC (difference in the abdominal circumference measured before and after colonoscopy) or directly evaluated by patients' perception (visual analogic scale, range 0-100). Results: 402 patients (44% male; mean age 57.7. \ub1. 12.5. years) received either HBB or placebo. No differences in ADR (31.7% vs. 28%, p=0.48), advanced-ADR (7.4% vs. 10.5%, p=0.35) were observed between HBB and placebo group, respectively. A significantly lower detection rate of flat/depressed lesions was observed in the HBB group (0.5% vs. 5.5%, p=0.003). The \u3b4AC and the bloating perception were comparable between the two groups (p=0.22 and p=0.48, respectively). Conclusions: HBB administered before colonoscope withdrawal does not increase adenoma detection rate and seems to hamper the visualization of flat/depressed lesions. This finding raises concerns on the indiscriminate use of HBB during colonoscopy. \ua9 2013 Editrice Gastroenterologica Italiana S.r.l