Differentiation of human bone marrow stem cells into cells with a neural phenotype: diverse effects of two specific treatments

BACKGROUND: It has recently been demonstrated that the fate of adult cells is not restricted to their tissues of origin. In particular, it has been shown that bone marrow stem cells can give rise to cells of different tissues, including neural cells, hepatocytes and myocytes, expanding their differentiation potential. RESULTS: In order to identify factors able to lead differentiation of stem cells towards cells of neural lineage, we isolated stromal cells from human adult bone marrow (BMSC). Cells were treated with: (1) TPA, forskolin, IBMX, FGF-1 or (2) retinoic acid and 2-mercaptoethanol (BME). Treatment (1) induced differentiation into neuron-like cells within 24 hours, while a longer treatment was required when using retinoic acid and BME. Morphological modifications were more dramatic after treatment (1) compared with treatment (2). In BMSC both treatments induced the expression of neural markers such as NF, GFAP, TUJ-1 and neuron-specific enolase. Moreover, the transcription factor Hes1 increased after both treatments. CONCLUSION: Our study may contribute towards the identification of mechanisms involved in the differentiation of stem cells towards cells of neural lineage

Duplications of KIAA1549 and BRAF screening by Droplet Digital PCR from formalin-fixed paraffin-embedded DNA is an accurate alternative for KIAA1549-BRAF fusion detection in pilocytic astrocytomas

Pilocytic astrocytomas represent the most common glioma subtype in young patients and account for 5.4% of all gliomas. They are characterized by alterations in the RAS–MAP kinase pathway, the most frequent being a tandem duplication on chromosome 7q34 involving the BRAF gene, resulting in oncogenic BRAF fusion proteins. BRAF fusion involving the KIAA1549 gene is a hallmark of pilocytic astrocytoma, but it has also been recorded in rare cases of gangliogliomas, 1p/19q co-deleted oligodendroglial tumors, and it is also a common feature of disseminated oligodendroglial-like leptomeningeal neoplasm. In some difficult cases, evidence for KIAA1549-BRAF fusion is of utmost importance for the diagnosis. Moreover, because the KIAA1549-BRAF fusion constitutively activates the MAP kinase pathway, it represents a target for drugs such as MEK inhibitors, and therefore, the detection of this genetic abnormality is highly relevant in the context of clinical trials applying such new approaches. In the present study, we aimed to use the high sensitivity of Droplet Digital PCR (DDPCR™) to predict KIAA1549-BRAF fusion on very small amounts of formalin-fixed paraffin-embedded tissue in routine practice. Therefore, we analyzed a training cohort of 55 pilocytic astrocytomas in which the KIAA1549-BRAF fusion status was known by RNA sequencing used as our gold standard technique. Then, we analyzed a prospective cohort of 40 pilocytic astrocytomas, 27 neuroepithelial tumors remaining difficult to classify (pilocytic astrocytoma versus ganglioglioma or diffuse glioma), 15 dysembryoplastic neuroepithelial tumors, and 18 gangliogliomas. We could demonstrate the usefulness and high accuracy (100% sensitivity and specificity when compared to RNA sequencing) of DDPCR™ to assess the KIAA1549-BRAF fusion from very low amounts of DNA isolated from formalin-fixed paraffin-embedded specimens. BRAF duplication is both necessary and sufficient to predict this fusion in most cases and we propose that this single analysis could be used in routine practice to save time, money, and precious tissue

The miR-139-5p regulates proliferation of supratentorial paediatric low-grade gliomas by targeting the PI3K/AKT/mTORC1 signalling

Paediatric low-grade gliomas (pLGGs) are a heterogeneous group of brain tumours associated with a high overall survival: however, they are prone to recur and supratentorial lesions are difficult to resect, being associated with high percentage of disease recurrence. Our aim was to shed light on the biology of pLGGs

CDX-2 expression correlates with clinical outcomes in MSI-H metastatic colorectal cancer patients receiving immune checkpoint inhibitors

Immune checkpoint inhibitors (ICIs) showed efficacy in metastatic colorectal cancer (mCRC) with mismatch-repair deficiency or high microsatellite instability (dMMR-MSI-H). Unfortunately, a patient's subgroup did not benefit from immunotherapy. Caudal-related homeobox transcription factor 2 (CDX-2) would seem to influence immunotherapy's sensitivity, promoting the chemokine (C-X-C motif) ligand 14 (CXCL14) expression. Therefore, we investigated CDX-2 role as a prognostic-predictive marker in patients with mCRC MSI-H. We retrospectively collected data from 14 MSI-H mCRC patients treated with ICIs between 2019 and 2021. The primary endpoint was the 12-month progression-free-survival (PFS) rate. The secondary endpoints were overall survival (OS), PFS, objective response rate (ORR), and disease control rate (DCR). The PFS rate at 12 months was 81% in CDX-2 positive patients vs 0% in CDX-2 negative patients (p = 0.0011). The median PFS was not reached (NR) in the CDX-2 positive group versus 2.07 months (95%CI 2.07-10.8) in CDX-2 negative patients (p = 0.0011). Median OS was NR in CDX-2-positive patients versus 2.17 months (95% Confidence Interval [CI] 2.17-18.7) in CDX2-negative patients (p = 0.026). All CDX-2-positive patients achieved a disease response, one of them a complete response. Among CDX-2-negative patients, one achieved stable disease, while the other progressed rapidly (ORR: 100% vs 0%, p = 0.0005; DCR: 100% vs 50%, p = 0.02). Twelve patients received 1st-line pembrolizumab (11 CDX-2 positive and 1 CDX-2 negative) not reaching median PFS, while two patients (1 CDX-2 positive and 1 CDX-2 negative) received 3rd-line pembrolizumab reaching a median PFS of 10.8 months (95% CI, 10.8-12.1; p = 0.036). Although our study reports results on a small population, the prognostic role of CDX-2 in CRC seems confirmed and could drive a promising predictive role in defining the population more sensitive to immunotherapy treatment. Modulating the CDX-2/CXCL14 axis in CDX-2-negative patients could help overcome primary resistance to immunotherapy

Chronic constipation diagnosis and treatment evaluation: The "CHRO.CO.DI.T.E." study

Background: According to Rome criteria, chronic constipation (CC) includes functional constipation (FC) and irritable bowel syndrome with constipation (IBS-C). Some patients do not meet these criteria (No Rome Constipation, NRC). The aim of the study was is to evaluate the various clinical presentation and management of FC, IBS-C and NRC in Italy. Methods: During a 2-month period, 52 Italian gastroenterologists recorded clinical data of FC, IBS-C and NRC patients, using Bristol scale, PAC-SYM and PAC-QoL questionnaires. In addition, gastroenterologists were also asked to record whether the patients were clinically assessed for CC for the first time or were in follow up. Diagnostic tests and prescribed therapies were also recorded. Results: Eight hundred seventy-eight consecutive CC patients (706 F) were enrolled (FC 62.5%, IBS-C 31.3%, NRC 6.2%). PAC-SYM and PAC-QoL scores were higher in IBS-C than in FC and NRC. 49.5% were at their first gastroenterological evaluation for CC. In 48.5% CC duration was longer than 10 years. A specialist consultation was requested in 31.6%, more frequently in IBS-C than in NRC. Digital rectal examination was performed in only 56.4%. Diagnostic tests were prescribed to 80.0%. Faecal calprotectin, thyroid tests, celiac serology, breath tests were more frequently suggested in IBS-C and anorectal manometry in FC. More than 90% had at least one treatment suggested on chronic constipation, most frequently dietary changes, macrogol and fibers. Antispasmodics and psychotherapy were more frequently prescribed in IBS-C, prucalopride and pelvic floor rehabilitation in FC. Conclusions: Patients with IBS-C reported more severe symptoms and worse quality of life than FC and NRC. Digital rectal examination was often not performed but at least one diagnostic test was prescribed to most patients. Colonoscopy and blood tests were the "first line" diagnostic tools. Macrogol was the most prescribed laxative, and prucalopride and pelvic floor rehabilitation represented a "second line" approach. Diagnostic tests and prescribed therapies increased by increasing CC severity

Existence of a Neutral-Impact Maxillo-Mandibular Displacement on Upper Airways Morphology

Current scientific evidence on how orthognathic surgery affects the airways morphology remains contradictory. The aim of this study is to investigate the existence and extension of a neutral-impact interval of bony segments displacement on the upper airways morphology. Its upper boundary would behave as a skeletal displacement threshold differentiating minor and major jaw repositioning, with impact on the planning of the individual case. Pre- and post-operative cone beam computed tomographies (CBCTs) of 45 patients who underwent maxillo-mandibular advancement or maxillary advancement/mandibular setback were analysed by means of a semi-automated three-dimensional (3D) method; 3D models of skull and airways were produced, the latter divided into the three pharyngeal subregions. The correlation between skeletal displacement, stacked surface area and volume was investigated. The displacement threshold was identified by setting three ∆Area percentage variations. No significant difference in area and volume emerged from the comparison of the two surgical procedures with bone repositioning below the threshold (approximated to +5 mm). A threshold ranging from +4.8 to +7 mm was identified, varying in relation to the three ∆Area percentages considered. The ∆Area increased linearly above the threshold, while showing no consistency in the interval ranging from −5 mm to +5 mm

Existence of a Neutral-Impact Maxillo-Mandibular Displacement on Upper Airways Morphology

Current scientific evidence on how orthognathic surgery affects the airways morphology remains contradictory. The aim of this study is to investigate the existence and extension of a neutral-impact interval of bony segments displacement on the upper airways morphology. Its upper boundary would behave as a skeletal displacement threshold differentiating minor and major jaw repositioning, with impact on the planning of the individual case. Pre- and post-operative cone beam computed tomographies (CBCTs) of 45 patients who underwent maxillo-mandibular advancement or maxillary advancement/mandibular setback were analysed by means of a semi-automated three-dimensional (3D) method; 3D models of skull and airways were produced, the latter divided into the three pharyngeal subregions. The correlation between skeletal displacement, stacked surface area and volume was investigated. The displacement threshold was identified by setting three ∆Area percentage variations. No significant difference in area and volume emerged from the comparison of the two surgical procedures with bone repositioning below the threshold (approximated to +5 mm). A threshold ranging from +4.8 to +7 mm was identified, varying in relation to the three ∆Area percentages considered. The ∆Area increased linearly above the threshold, while showing no consistency in the interval ranging from −5 mm to +5 mm

Analysis of N-ras gene mutations in medulloblastomas by polymerase chain reaction and oligonucleotide probes in formalin-fixed, paraffin-embedded tissues.

Precise data on the incidence of transforming ras oncogenes in pediatric tumors and the correlations with the histopathological properties of the tumors are very limited. Additionally the presence of ras activation in medulloblastomas has not been investigated so far. Using a combination of techniques including in vitro gene amplification by polymerase chain reaction (PCR) and detection of single base mutations by sequence-specific oligonucleotides we studied N-ras activation (mutations at codon 12, 13, and 61) in 32 medulloblastomas. DNA was isolated from 20-mu-m sections of formalin-fixed paraffin-embedded tissue. Mutations were found in 3 out of 32 examined medulloblastomas. In all cases only mutations of codon 61 were found: two of three mutations were C to A mutations at position 1 of the codon 61 (leading to a substitution of a glutamine residue for a lysine) and one was A to T mutation at position 3 in the same codon (glutamine-histidine). Our results indicate 10% incidence N-ras mutation in medulloblastoma, higher than in other CNS tumors studied so far. The main advantages of the procedure described are its greatly improved sensitivity, the increased speed with which tumor samples can be analyzed, and the possibility of using paraffin-embedded sections to analyze various rare tumors in retrospect