Stefano Avanzini

Induction of RET Dependent and Independent Pro-Inflammatory Programs in Human Peripheral Blood Mononuclear Cells from Hirschsprung Patients

Hirschsprung disease (HSCR) is a rare congenital anomaly characterized by the absence of enteric ganglia in the distal intestinal tract. While classified as a multigenic disorder, the altered function of the RET tyrosine kinase receptor is responsible for the majority of the pathogenesis of HSCR. Recent evidence demonstrate a strong association between RET and the homeostasis of immune system. Here, we utilize a unique cohort of fifty HSCR patients to fully characterize the expression of RET receptor on both innate (monocytes and Natural Killer lymphocytes) and adaptive (B and T lymphocytes) human peripheral blood mononuclear cells (PBMCs) and to explore the role of RET signaling in the immune system. We show that the increased expression of RET receptor on immune cell subsets from HSCR individuals correlates with the presence of loss-of-function RET mutations. Moreover, we demonstrate that the engagement of RET on PBMCs induces the modulation of several inflammatory genes. In particular, RET stimulation with glial-cell line derived neurotrophic factor family (GDNF) and glycosyl-phosphatidylinositol membrane anchored co-receptor α1 (GFRα1) trigger the up-modulation of genes encoding either for chemokines (CCL20, CCL2, CCL3, CCL4, CCL7, CXCL1) and cytokines (IL-1β, IL-6 and IL-8) and the down-regulation of chemokine/cytokine receptors (CCR2 and IL8-Rα). Although at different levels, the modulation of these “RET-dependent genes” occurs in both healthy donors and HSCR patients. We also describe another set of genes that, independently from RET stimulation, are differently regulated in healthy donors versus HSCR patients. Among these “RET-independent genes”, there are CSF-1R, IL1-R1, IL1-R2 and TGFβ-1, whose levels of transcripts were lower in HSCR patients compared to healthy donors, thus suggesting aberrancies of inflammatory responses at mucosal level. Overall our results demonstrate that immune system actively participates in the physiopathology of HSCR disease by modulating inflammatory programs that are either dependent or independent from RET signaling.

Genomics approach to the analysis of bacterial communities dynamics in Hirschsprung’s disease-associated enterocolitis: a pilot study

IntroductionThe most invalidating and life-threatening complication in Hirschsprung’s disease patients (HSCR) is Hirschsprung’s disease-associated enterocolitis (HAEC). The mechanisms underlying enterocolitis have not been identified. The limited knowledge of the role of intestinal microflora is in part due to the complexity of the intestinal microbiome and to the limitation of cultivation-based technologies, given that less than 25% of the intestinal bacterial species can be cultured.Materials and methodsWe used amplified ribosomal DNA restriction analysis (ARDRA) with four different restriction enzymes to study variations of microflora composition of the stools of a selected HSCR patient in different clinical conditions (acute phase vs. remission).ResultsWe assessed a total of 15 stool specimens belonging to the same 3-year-old male patient suffering from HSCR, which were harvested during 4 HAEC episodes and remission phases. Restriction analysis showed that HAEC episodes seem to cluster together at ARDRA analysis, thus suggesting a sort of predisposing bacterial community for HAEC development and the need for a microflora equilibrium to maintain wellness.ConclusionsThis approach proved to be effective, useful and powerful in assessing microflora dynamics and indicated that the differences in microflora associated with acute HAEC or remission are likely to result from a combination of disease activity and different antibiotic therapies. ARDRA proved to be useful in discriminating disease versus remission. Our findings indicated that HAEC results from a change in the equilibrium between bacterial species or from altered discrimination of harmless from harmful microorganisms, challenging the definition of pathogenic and non-pathogenic species. Based on these results, we propose ARDRA as a rapid inexpensive tool to assess microflora dynamics during HAEC episodes.

Redo surgery in Hirschsprung disease: what did we learn? Unicentric experience on 70 patients

PURPOSE This article is aimed at describing a series of 70 patients who underwent a redo pull-through in the period between January 1991 and January 2007 and to compare them with a series of consecutive patients with Hirschsprung disease who underwent a single pull-through. METHODS The notes were reviewed, and a comprehensive interview based on a disease-specific questionnaire for continence and quality of life was submitted to all eligible patients. We used a number of consecutive patients who underwent a single pull-through for Hirschsprung disease as control group. RESULTS Seventy patients were included. Residual aganglionosis in pulled-through bowel or retained aganglionic rectum represented the indications to reoperation in most cases. Intestinal obstruction represented the leading symptom in more than 60% of patients. We adopted either the endorectal pull-through, the Duhamel, or the Swenson procedure. Thirty-six patients were assessed for long-term outcome. One third experienced complications. Long-term continence was satisfactory in 70%. Enterocolitis, soiling, and perineal excoriations were complained by 30%. Excellent to good perspectives were described by 94%, and excellent to good cosmetic results by 22%. The only significant difference with control group (109 patients) involved cosmetic appearance, whereas soiling, although more frequent, could not be considered significantly increased. CONCLUSIONS Patients who experience failure of a pull-through must go through multiple procedures to achieve definitive results. Although overall outcome does not significantly differ from that of a single effective pull-through, great efforts should be made to minimize complications and avoid the need for a redo, which requires experienced surgeons and highly committed families for a longer course of the disease.

Simultaneous occurrence of pancreatoblastoma and neuroblastoma in a newborn with beckwith-wiedemann syndrome.

Beckwith-Wiedemann syndrome is associated with an increased risk of tumors in the pediatric age. We report the case of a newborn with Beckwith-Wiedemann syndrome presenting the simultaneous occurrence of a cystic pancreatoblastoma and an adrenal neuroblastoma. Surgery was required to treat the pancreatoblastoma, and a “wait and see” policy was adopted for the neuroblastoma, which spontaneously regressed within a few months.

Image-defined risk factors in unresectable neuroblastoma: SIOPEN study on incidence, chemotherapy-induced variation, and impact on surgical outcomes

To evaluate the impact of image‐defined risk factor (IDRF) modification after chemotherapy on surgical outcomes, event‐free survival (EFS), and overall survival (OS) among patients enrolled in the European Unresectable Neuroblastoma (EUNB) study.

Laparoscopic Proximal Roux-en-Y Gastrojejunal Diversion in Children: Preliminary Experience from a Single Center

BACKGROUND Neurologically impaired children (NIC) have a high risk of recurrence of gastroesophageal reflux (GER) following fundoplication. A postpyloric feeding tube may be useful when gastric emptying disorders occur; however, dislocation and difficulty in feeding management often require more aggressive procedures. Total esophagogastric dissociation (Bianchis TEGD) is an alternative to the classic fundoplication procedure, whereas laparoscopic gastric bypass is a frequently performed procedure in morbid obesity, improving gastric outlet. AIM The aim of this paper is to present a preliminary experience on the laparoscopic Roux-en-Y gastrojejunal bypass, associated with Nissen fundoplication and gastrostomy, to treat and prevent GER in NIC with gastric emptying disorders. MATERIALS AND METHODS Eight neurologically impaired children underwent surgical treatment because of feeding problems and pulmonary complications. The procedure included: 1) hiatoplasty, 2) Nissen fundoplication, 3) 20-cm Roux-en-Y gastrojejunal anastomosis and jejuno-jejunal anastomosis, and 4) gastrostomy. RESULTS All cases were fed on postoperative day 3 without any intraoperative complications. One case developed an obstruction of the distal anastomosis due to adhesion and needed reoperation. Outcome was clinically evaluated with serial upper gastrointestinal contrast studies and endoscopies. CONCLUSIONS Laparoscopic proximal Roux-en-Y gastrojejunal diversion, without gastric resection, is a safe, feasible procedure that improves gastric emptying and reduces the risk of GER recurrence. Yet, long-term results still have to be evaluated.

Brachiocephalic vein for percutaneous ultrasound-guided central line positioning in children: A 20-month preliminary experience with 109 procedures

Ultrasound‐guided (USG) cannulation of the brachiocephalic vein (BCV) is gaining worldwide consensus for central venous access in children. This study reports a 20‐month experience with this approach in children.

Life-threatening bilateral adrenal cystic neuroblastoma in an infant.

A case of neonatal bilateral adrenal mass causing severe respiratory distress and requiring an emergency debulking surgical procedure is reported. Histopathology revealed a cystic neuroblastoma stroma poor, poorly differentiated, without MYCN amplification and 1p deletion. During postoperative follow-up, a progression to stage 4s was observed, characterized by liver involvement. According to good prognostic indexes, no further treatment was administered. Both adrenal masses and hepatic nodules showed progressive decrease in size, till complete disappearance. The authors encourage a multidisciplinary approach to develop the best patient-related strategy for cystic neuroblastoma, thus reducing complications rate.

Use of failure modes, effects, and criticality analysis to compare the vulnerabilities of laparoscopic versus open appendectomy

Purpose To measure the feasibility of using FMECA applied to the surgery and then compare the vulnerabilities of laparoscopic versus open appendectomy by using FMECA. Methods The FMECA study was performed on each single selected phase of appendectomy and on complication-related data during the period January 1, 2009, to December 31, 2010. The risk analysis phase was completed by evaluation of the criticality index (CI) of each appendectomy-related failure mode (FM). The CI is calculated by multiplying the estimated frequency of occurrence (O) of the FM, by the expected severity of the injury to the patient (S), and the detectability (D) of the FM. Results In the first year of analysis (2009), 177 appendectomies were performed, 110 open and 67 laparoscopic. Eleven adverse events were related to the open appendectomy: 1 bleeding (CI: 8) and 10 postoperative infections (CI: 32). Three adverse events related to the laparoscopic approach were recorded: 1 postoperative infection (CI: 8) and 2 incorrect extractions of the appendix through the umbilical port (CI: 6). In the second year of analysis (2010), 158 appendectomies were performed, 69 open and 89 laparoscopic. Four adverse events were related to the open appendectomy: 1 incorrect management of the histological specimen (CI: 2), 1 dehiscence of the surgical wound (CI: 6), and 2 infections (CI: 6). No adverse events were recorded in laparoscopic approach. Conclusion FMECA helped the staff compare the 2 approaches through an accurate step-by-step analysis, highlighting that laparoscopic appendectomy is feasible and safe, associated with a lower incidence of infection and other complications, reduced length of hospital stay, and an apparent lower procedure-related risk.

Management of nontuberculous mycobacterial lymphadenitis in a tertiary care children's hospital: A 20 year experience

PURPOSE Nontuberculous mycobacteria are uncommon cause of chronic cervicofacial lymphadenitis in healthy children. We describe clinical features and management strategies of cervicofacial nontuberculous mycobacterium lymphadenitis in a tertiary pediatric hospital. METHODS Retrospective analysis of medical records of children discharged from 1992 to 2014 with a diagnosis of cervicofacial nontuberculous mycobacterium was made. Diagnosis certainty was based on microhistological investigations. Clinical stage was evaluated according to lymph node size and presence of fistulas. Successful therapy was defined by the regression of the lymph node enlargement (>75%) or complete surgical excision without relapse. RESULTS Cervicofacial nontuberculous mycobacterium was diagnosed in 33 patients. Complete excision was performed in 73% of cases primarily observed in our hospital, while 83% of those referred from other hospitals required further surgical treatment. No case of relapse was observed after one year of follow-up. CONCLUSIONS We recommend surgical approach as the first therapeutic option in the management of cervicofacial nontuberculous mycobacterium lymphadenitis. LEVELS OF EVIDENCE Prognosis and Retrospective Study - Level II.