Bidirectional Approach with PIPAC and Systemic Chemotherapy for Patients with Synchronous Gastric Cancer Peritoneal Metastases (GCPM).

BACKGROUND: This study evaluated the efficacy of pressurized intraperitoneal aerosol chemotherapy (PIPAC) with systemic chemotherapy as a bidirectional approach for gastric cancer (GC) patients with synchronous peritoneal metastases (SPM). METHODS: A retrospective analysis of a prospective PIPAC database was queried for patients who underwent a bidirectional approach between October 2019 and April 2022 at two high-volume GC surgery units in Italy (Verona and Siena). Surgical and oncological outcomes were analyzed. RESULTS: Between October 2019 and April 2022, 74 PIPAC procedures in 42 consecutive patients with Eastern Cooperative Oncology Group performance status ≤2 were performed-32 patients treated in Verona and 10 in Siena. Twenty-seven patients (64%) were female and median age at first PIPAC was 60.5 years (I-III quartiles: 49-68 years). Median Peritoneal Cancer Index (PCI) was 16 (I-III quartiles: 8-26) and 25 patients (59%) had at least two PIPAC procedures. Major complications according to the Common Terminology Criteria for Adverse Events (CTCAE; 3 and 4) occurred in three (4%) procedures, and, according to the Clavien-Dindo classification (>3a), one (1%) severe complication occurred. There were no reoperations or deaths within 30 days. Median overall survival (mOS) from diagnosis was 19.6 months (range 14-24), and mOS from first PIPAC was 10.5 months (range 7-13). Excluding cases with very heavy metastatic peritoneal burden, with PCI from 2 to 26, treated with more than one PIPAC, mOS from diagnosis was 22 months (range 14-39). Eleven patients (26%) underwent curative-intent surgery after a bidirectional approach. R0 was achieved in nine (82%) patients and complete pathological response was obtained in three (27%) cases. CONCLUSIONS: Patient selection is associated with bidirectional approach efficacy and feasibility for SPM GC treatment, which may allow potentially curative surgical radicalization in highly selected cases.

Assessing pain in children with autism spectrum disorders: findings from a preliminary validation study.

AIMS: Assessing pain in children with autism spectrum disorders (ASDs) can be extremely challenging, since many cannot self-report pain. This study aims to test the validity of the Non-Communicating Children's Pain Checklist - Revised (NCCPC-R) in identifying pain in children and adolescents affected by ASDs. MATERIALS AND METHODS: A two-phase validation study based on (a) the translation and cultural adaptation of the NCCPC-R to Italian and to ASD-specific needs and context; and (b) the validation of a modified, 32-item version of the NCCPC-R. In all, 141 carers of children aged 6-16 years with ASDs were asked to recall an in-pain episode and a not-in-pain episode of their child and to rate on a 3-point scale (0 = not at all, 3 = very often) each behaviour included in the tool. Internal consistency (Cronbach's α), explorative and confirmative factorial structure, as well as concurrent and discriminant validity, were all assessed. RESULTS: Confirmatory factor analysis established the revised version of the NCCPC-R for children with ASDs (named = NCCPC-RASD ), formed from 10 of the original 30 items categorised into three factors ('Changing in mood', 'Increasing in tension' and 'Alerting reaction') to have an acceptable level of reliability. The tool was internally consistent (α = 0.741 during in-pain episodes, α = 0.790 during not-in-pain episodes) and was able to discriminate between in-pain episodes (13.36 out of 40; CI 95% 12.34-14.39) and not-in-pain episodes (7.84 out of 40; CI 95% 6.86-8.82, p < 0.001). CONCLUSIONS: These results provide preliminary evidence that the 10-item version of the NCCPC-RASD is a reliable and valid tool for assessing pain in children with ASD.

Mutations of cytochrome c identified in patients with thrombocytopenia THC4 affect both apoptosis and cellular bioenergetics.

Inherited thrombocytopenias are heterogeneous diseases caused by at least 20 genes playing different role in the processes of megakaryopoiesis and platelet production. Some forms, such as thrombocytopenia 4 (THC4), are very rare and not well characterized. THC4 is an autosomal dominant mild thrombocytopenia described in only one large family from New Zealand and due to a mutation (G41S) of the somatic isoform of the cytochrome c (CYCS) gene. We report a novel CYCS mutation (Y48H) in patients from an Italian family. Similar to individuals carrying G41S, they have platelets of normal size and morphology, which are only partially reduced in number, but no prolonged bleeding episodes. In order to determine the pathogenetic consequences of Y48H, we studied the effects of the two CYCS mutations in yeast and mouse cellular models. In both cases, we found reduction of respiratory level and increased apoptotic rate, supporting the pathogenetic role of CYCS in thrombocytopenia.

Children of a lesser god or miracles? An emotional and behavioural profile of children born to mothers on dialysis in Italy: a multicentre nationwide study 2000-12.

BACKGROUND: Pregnancy on dialysis is increasingly being reported. This study evaluates the behavioural profile of the children of mothers on dialysis and the parental stress their mothers undergo when compared with a group of mothers affected by a different chronic disease (microcythaemia) and a group of healthy control mothers. METHODS: Between 2000 and 2012, 23 on-dialysis mothers gave birth to 24 live-born children in Italy (23 pregnancies, 1 twin pregnancy, one of the twins deceased soon after delivery); of these, 16 mothers and 1 father (whose wife died before the inquiry) were included in the study (1 mother had died and the father was unavailable; 2 were not asked to participate because their children had died and 3 were unavailable; children: median age: 8.5, min-max: 2-13 years). Twenty-three mothers affected by transfusion-dependent microcythaemia or drepanocitosis (31 pregnancies, 32 children) and 35 healthy mothers (35 pregnancies, 35 children; median age of the children: 7, min-max: 1-13 years) were recruited as controls. All filled in the validated questionnaires: 'Child Behaviour Checklist' (CBCL) and the 'Parental Stress Index-Short Form' (PSI-SF). RESULTS: The results of the CBCL questionnaire were similar for mothers on dialysis and healthy controls except for pervasive developmental problems, which were significantly higher in the dialysis group, while microcythaemia mothers reported higher emotional and behavioural problems in their children in 8 CBCL sub-scales. Two/16 children in the dialysis and 3/32 in the microcythaemia group had pathological profiles, as assessed by T-scores (p: ns). PSI-SF indicated a normal degree of parental stress in microcythaemia subjects and healthy controls, while mothers on dialysis declared significantly lower stress, suggesting a defensive response in order to minimize problems, stress or negativity in their relationship with their child. CONCLUSIONS: According to the present analysis, the emotional and behavioural outcome is normal in most of the children from on-dialysis mothers. A 'positive defence' in the dialysis mothers should be kept in mind when tailoring psychological support for this medical miracle.

Intact cell mass spectrometry coupled with machine learning reveals minute changes induced by single gene silencing.

Intact (whole) cell MALDI TOF mass spectrometry is a commonly used tool in clinical microbiology for several decades. Recently it was introduced to analysis of eukaryotic cells, including cancer and stem cells. Besides targeted metabolomic and proteomic applications, the intact cell MALDI TOF mass spectrometry provides a sufficient sensitivity and specificity to discriminate cell types, isogenous cell lines or even the metabolic states. This makes the intact cell MALDI TOF mass spectrometry a promising tool for quality control in advanced cell cultures with a potential to reveal batch-to-batch variation, aberrant clones, or unwanted shifts in cell phenotype. However, cellular alterations induced by change in expression of a single gene has not been addressed by intact cell mass spectrometry yet. In this work we used a well-characterized human ovarian cancer cell line SKOV3 with silenced expression of a tumor suppressor candidate 3 gene (TUSC3). TUSC3 is involved in co-translational N-glycosylation of proteins with well-known global impact on cell phenotype. Altogether, this experimental design represents a highly suitable model for optimization of intact cell mass spectrometry and analysis of spectral data. Here we investigated five machine learning algorithms (k-nearest neighbors, decision tree, random forest, partial least squares discrimination, and artificial neural network) and optimized their performance either in pure populations or in two-component mixtures composed of cells with normal or silenced expression of TUSC3. All five algorithms reached accuracy over 90 % and were able to reveal even subtle changes in mass spectra corresponding to alterations of TUSC3 expression. In summary, we demonstrate that spectral fingerprints generated by intact cell MALDI-TOF mass spectrometry coupled to a machine learning classifier can reveal minute changes induced by alteration of a single gene, and therefore contribute to the portfolio of quality control applications in routine cell and tissue cultures.

Characterization of a proteasome and TAP-independent presentation of intracellular epitopes by HLA-B27 molecules.

Nascent HLA-class I molecules are stabilized by proteasome-derived peptides in the ER and the new complexes proceed to the cell surface through the post-ER vesicles. It has been shown, however, that less stable complexes can exchange peptides in the Trans Golgi Network (TGN). HLA-B27 are the most studied HLA-class I molecules due to their association with Ankylosing Spondylitis (AS). Chimeric proteins driven by TAT of HIV have been exploited by us to deliver viral epitopes, whose cross-presentation by the HLA-B27 molecules was proteasome and TAP-independent and not restricted to Antigen-Presenting Cells (APC). Here, using these chimeric proteins as epitope suppliers, we compared with each other and with the HLA-A2 molecules, the two HLA-B*2705 and B*2709 alleles differing at residue 116 (D116H) and differentially associated with AS. We found that the antigen presentation by the two HLA-B27 molecules was proteasome-, TAP-, and APC-independent whereas the presentation by the HLA-A2 molecules required proteasome, TAP and professional APC. Assuming that such difference could be due to the unpaired, highly reactive Cys-67 distinguishing the HLA-B27 molecules, C67S mutants in HLA-B*2705 and B*2709 and V67C mutant in HLA-A*0201 were also analyzed. The results showed that this mutation did not influence the HLA-A2-restricted antigen presentation while it drastically affected the HLA-B27-restricted presentation with, however, remarkable differences between B*2705 and B*2709. The data, together with the occurrence on the cell surface of unfolded molecules in the case of C67S-B*2705 mutant but not in that of C67S-B*2709 mutant, indicates that Cys-67 has a more critical role in stabilizing the B*2705 rather than the B*2709 complexes.

Recessive germline SDHA and SDHB mutations causing leukodystrophy and isolated mitochondrial complex II deficiency.

BACKGROUND: Isolated complex II deficiency is a rare form of mitochondrial disease, accounting for approximately 2% of all respiratory chain deficiency diagnoses. The succinate dehydrogenase (SDH) genes (SDHA, SDHB, SDHC and SDHD) are autosomally-encoded and transcribe the conjugated heterotetramers of complex II via the action of two known assembly factors (SDHAF1 and SDHAF2). Only a handful of reports describe inherited SDH gene defects as a cause of paediatric mitochondrial disease, involving either SDHA (Leigh syndrome, cardiomyopathy) or SDHAF1 (infantile leukoencephalopathy). However, all four SDH genes, together with SDHAF2, have known tumour suppressor functions, with numerous germline and somatic mutations reported in association with hereditary cancer syndromes, including paraganglioma and pheochromocytoma. METHODS AND RESULTS: Here, we report the clinical and molecular investigations of two patients with histochemical and biochemical evidence of a severe, isolated complex II deficiency due to novel SDH gene mutations; the first patient presented with cardiomyopathy and leukodystrophy due to compound heterozygous p.Thr508Ile and p.Ser509Leu SDHA mutations, while the second patient presented with hypotonia and leukodystrophy with elevated brain succinate demonstrated by MR spectroscopy due to a novel, homozygous p.Asp48Val SDHB mutation. Western blotting and BN-PAGE studies confirmed decreased steady-state levels of the relevant SDH subunits and impairment of complex II assembly. Evidence from yeast complementation studies provided additional support for pathogenicity of the SDHB mutation. CONCLUSIONS: Our report represents the first example of SDHB mutation as a cause of inherited mitochondrial respiratory chain disease and extends the SDHA mutation spectrum in patients with isolated complex II deficiency.

Uninsured immigrant and refugee children presenting to Canadian paediatric emergency departments: Disparities in help-seeking and service delivery.

INTRODUCTION: Access to health care for medically uninsured immigrant and refugee children is a public health concern due to the consequences of delayed or substandard care for child development and health. OBJECTIVE: To explore possible differences in help-seeking and service delivery across migratory statuses, institutions and provinces. METHODS: A review was undertaken of 2035 emergency files of immigrant, refugee and undocumented children without provincial health care coverage who sought care at three major paediatric hospitals in Montreal (Quebec) and Toronto (Ontario) during 2008 and 2009. RESULTS: Refugee claimant children with Interim Federal Health Program benefits consulted for less urgent problems than the overall hospital population, except in one hospital that had a multicultural paediatric ambulatory clinic. Undocumented children and new permanent resident immigrant children within the three-month waiting period for provincial health care coverage were over-represented in the very urgent triage category and presented more often for injuries, trauma and mental health problems than did refugee claimant children. DISCUSSION/CONCLUSIONS: Wide interhospital differences suggest that the predicament of limited access to health care of these groups of vulnerable medically uninsured children needs to be addressed through further research to inform policies and develop training.

INTRODUCTION: L'accès aux soins de santé pour les enfants immigrants et réfugiés qui n'ont pas d'assurance est un problème de santé publique, en raison des conséquences du retard des soins ou des soins de second ordre sur leur développement et leur santé. OBJECTIF: Explorer les différences possibles de recherche d'aide et de prestation des soins selon les statuts migratoires, les établissements et les provinces. MÉTHODOLOGIE: Les chercheurs ont entrepris une analyse de 2 035 dossiers urgents d'enfants immigrants, réfugiés et sans papiers, sans assurance-maladie provinciale, qui ont consulté dans trois grands hôpitaux pédiatriques de Montréal, au Québec, et de Toronto, en Ontario, en 2008 et en 2009. RÉSULTATS: Les enfants demandeurs du statut de réfugié profitant des prestations du Programme fédéral de santé intérimaire consultaient pour des problèmes moins urgents que l'ensemble de la population hospitalière, sauf dans un hôpital doté d'une clinique ambulatoire pédiatrique multiculturelle. Les enfants sans papiers et les enfants immigrants qui étaient de nouveaux résidents permanents assujettis à la période d'attente de trois mois avant d'avoir droit à l'assurance-maladie provinciale étaient surreprésentés dans la catégorie de triage très urgent et présentaient plus souvent des blessures, des traumatismes et des problèmes de santé mentale que les enfants demandeurs du statut de réfugié. EXPOSÉ ET CONCLUSIONS: D'après les vastes différences interhos-pitalières, il faudrait poursuivre les recherches sur la situation difficile causée par l'accès limité aux soins de santé de ces groupes d'enfants non assurés vulnérables pour étayer les politiques et élaborer les formations.

A novel heteroleptic Cu(II)-phenanthroline-UDCA complex as lipoxygenase inhibitor and ER-stress inducer in cancer cell lines.

A new heteroleptic copper(II) compound named C0-UDCA was prepared by reaction of [Cu(phen)2(OH2)](ClO4)2 (C0) with the bile ursodeoxycholic acid (UDCA). The resulting compound is able to inhibit the lipoxygenase enzyme showing more efficacy than the precursors C0 and UDCA. Molecular docking simulations clarified the interactions with the enzyme as due to allosteric modulation. The new complex shows antitumoral effect on ovarian (SKOV-3) and pancreatic (PANC-1) cancer cells at the Endoplasmic Reticulum (ER) level by activating the Unfolded Protein Response. In particular, the chaperone BiP, the pro-apoptotic protein CHOP and the transcription factor ATF6 are upregulated in the presence of C0-UDCA. The combination of Intact Cell MALDI-MS and statistical analysis have allowed us to discriminate between untreated and treated cells based on their mass spectrometry fingerprints.