Biochemical characterization on muscle tissue of a novel biallelic ACO2 mutation in an infant with progressive encephalopathy.

The ACO2 gene encodes the mitochondrial protein aconitate hydratase, which is responsible for catalyzing the interconversion of citrate into isocitrate in the tricarboxylic acid (TCA) cycle. Mitochondrial aconitase is expressed ubiquitously, and deficiencies in TCA-cycle enzymes have been reported to cause various neurodegenerative diseases due to disruption of cellular energy metabolism and development of oxidative stress. We investigated a severe early infantile-onset neurometabolic syndrome due to a homozygous novel variant in exon 13 of the ACO2 gene. The in vitro pathogenicity of this variant of unknown significance was demonstrated by the loss of both protein expression and its enzymatic activity on muscle tissue sample taken from the patient. The patient presented with progressive encephalopathy soon after birth, characterized by hypotonia, progressive severe muscle atrophy, and respiratory failure. Serial brain magnetic resonance imaging showed progressive abnormalities compatible with a metabolic disorder, possibly mitochondrial. Muscle biopsy disclosed moderate myopathic alterations and features consistent with a mitochondriopathy albeit nonspecific. The course was characterized by progressive worsening of the clinical and neurological picture, and the patient died at 5 months of age. This study provides the first report on the validation in muscle from human subjects regarding in vitro analysis for mitochondrial aconitase activity. To our knowledge, no prior reports have demonstrated a correlation of phenotypic and diagnostic characteristics with in vitro muscle enzymatic activity of mitochondrial aconitase in humans. In conclusion, this case further expands the genetic spectrum of ACO2 variants and defines a complex case of severe neonatal neurometabolic disorder.

Neurocognitive and Psychological Outcomes among Children and Adolescents with Brain Tumors: Development of an Observational and Longitudinal Prospective Study Protocol.

Children and adolescents affected by brain tumors are at risk for neuropsychological sequelae that need to be evaluated in order to plan adequate rehabilitation programs, and to support their development and recovery. This work aims to describe an innovative prospective observational study protocol for the early evaluation and monitoring over time of neuropsychological outcomes in this pediatric population. Pediatric patients aged 3-17 with a brain tumor diagnosis will be assessed through the use of a battery of Italian standardized neuropsychological tests, with good psychometric properties and age-appropiate, at three different time points of their clinical course: at diagnosis and before surgery (T0), after surgical removal and before the start of potential adjuvant therapies (T1), and at the one-year follow-up after potential adjuvant therapies (T2). This study will allow clinicians to support the neuropsychological development of these children by promoting appropriate and timely rehabilitation and educational programs from the early phases of their clinical course.

A Schematic Approach to Defining the Prevalence of COL VI Variants in Five Years of Next-Generation Sequencing.

OBJECTIVE: To define the prevalence of variants in collagen VI genes through a next-generation sequencing (NGS) approach in undiagnosed patients with suspected neuromuscular disease and to propose a diagnostic flowchart to assess the real pathogenicity of those variants. METHODS: In the past five years, we have collected clinical and molecular information on 512 patients with neuromuscular symptoms referred to our center. To pinpoint variants in COLVI genes and corroborate their real pathogenicity, we sketched a multistep flowchart, taking into consideration the bioinformatic weight of the gene variants, their correlation with clinical manifestations and possible effects on protein stability and expression. RESULTS: In Step I, we identified variants in COLVI-related genes in 48 patients, of which three were homozygous variants (Group 1). Then, we sorted variants according to their CADD score, clinical data and complementary studies (such as muscle and skin biopsy, study of expression of COLVI on fibroblast or muscle and muscle magnetic resonance). We finally assessed how potentially pathogenic variants (two biallelic and 12 monoallelic) destabilize COL6A1-A2-A3 subunits. Overall, 15 out of 512 patients were prioritized according to this pipeline. In seven of them, we confirmed reduced or absent immunocytochemical expression of collagen VI in cultured skin fibroblasts or in muscle tissue. CONCLUSIONS: In a real-world diagnostic scenario applied to heterogeneous neuromuscular conditions, a multistep integration of clinical and molecular data allowed the identification of about 3% of those patients harboring pathogenetic collagen VI variants.

Juvenile Myasthenia Gravis in a 14-year-old adolescent masked by mood disorder: the complex balance between neurology and psychiatry.

Juvenile Myasthenia Gravis (JMG) is a neuromuscular disease, often characterized at onset by fatigue and fluctuating weakness. We report a case of a girl affected by severe mood disorder, in which the diagnosis of JMG and its treatment were challenged by the concomitant psychiatric condition. A 14-year-old girl, with a history of severe mood disorder and emotional dysregulation, had been treated with benzodiazepines, sertraline, and antipsychotics, reporting generalized fatigability, weakness, and drowsiness, first ascribed to her psychiatric condition and therapy. After a suicide attempt, she was hospitalized and a neurological assessment revealed a fluctuating ptosis and facial weakness, that improved with rest. The diagnosis of JMG was confirmed by repeated nerve stimulation test, and by the response to pyridostigmine. Antibodies anti-AChR and anti-MuSK were negative. JMG diagnosis may be harder in adolescents with psychiatric comorbidity. Moreover, the neurological condition limits the choice of the appropriate psychopharmacotherapy.

Gut Microbiota and Associated Mucosal Immune Response in Eosinophilic Granulomatosis with Polyangiitis (EGPA).

Eosinophilic granulomatosis with polyangiitis (EGPA) is an anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis. A genome-wide association study showed a correlation between ANCA-negative EGPA and variants of genes encoding proteins with intestinal barrier functions, suggesting that modifications of the mucosal layer and consequent gut dysbiosis might be involved in EGPA pathogenesis. Here, we characterized the gut microbiota (GM) composition and the intestinal immune response in a cohort of EGPA patients. Faeces from 29 patients and 9 unrelated healthy cohabitants were collected, and GM and derived metabolites' composition were compared. Seven intestinal biopsies from EGPA patients with gastrointestinal manifestations were analysed to assess the T-cell distribution and its correlation with GM and EGPA clinical and laboratory features. No significant differences in GM composition, nor in the total amount of faecal metabolites, emerged between patients and controls. Nevertheless, differences in bacterial taxa abundances and compositional GM-derived metabolites profile were observed. Notably, an enrichment of potential pathobionts (Enterobacteriacee and Streptococcaceae) was found in EGPA, particularly in patients with active disease, while lower levels were found in patients on immunosuppression, compared with non-immunosuppressed ones. Significantly lower amounts of hexanoic acid were found in patients, compared to controls. The analysis of the immune response in the gut mucosa revealed a high frequency of IFN-γ/IL-17-producing T lymphocytes, and a positive correlation between EGPA disease activity and intestinal T-cell levels. Our data suggest that an enrichment in potential intestinal pathobionts might drive an imbalanced inflammatory response in EGPA.

Diagnosis and treatment of Chiari malformation type 1 in children: the International Consensus Document.

BACKGROUND: Chiari malformation type 1 (CM1) is a rare condition where agreed classification and treatment are still missing. The goal of this study is to achieve a consensus on the diagnosis and treatment of CM1 in children. METHODS: A multidisciplinary panel formulated 57 provisional statements based on a review of the literature. Thirty-four international experts (IE) participated in a Delphi study by independently rating each statement on a 4-point Likert scale ("strongly disagree," "disagree," "agree," "strongly agree"). Statements that were endorsed ("agree" or "strongly agree") by < 75% of raters were re-formulated, or new statements were added, and another Delphi round followed (up to a maximum of three). RESULTS: Thirty-five IE were contacted and 34 agreed to participate. A consensus was reached on 30/57 statements (52.6%) after round 1. Three statements were added, and one removed. After round 2, agreement was reached on 56/59 statements (94.9%). Finally, after round 3, which took place during the 2019 Chiari Consensus Conference (Milan, Italy), agreement was reached on 58/59 statements (98.3%) about four main sections (Definition and Classification, Planning, Surgery, Isolated Syringomyelia). Only one statement did not gain a consensus, which is the "definition of radiological failure 24 month post-surgery." CONCLUSIONS: The consensus document consists of 58 statements (24 on diagnosis, 34 on treatment), serving clinicians and researchers following children with CM1. There is a clear need for establishing an international network and registry and to promote collaborative studies to increase the evidence base and optimize the long-term care of this patient population.

Myeloid neoplasms and autoimmune diseases: markers of association.

OBJECTIVES: To investigate the prognostic significance of concomitant autoimmune diseases (ADs) in myeloproliferative neoplasms (MPNs). METHODS: 435 subjects with a diagnosis of MPNs were included in this observational single institution longitudinal study. Of them, 34 patients presented an overt AD at diagnosis of MPN. Clinical presenting features, progression-free and overall survival were compared between MPN subgroups in relation to co-existence of AD at diagnosis of MPN. RESULTS: Compared to cases without ADs, the subjects with ADs were significantly younger, had lower haemoglobin and haematocrit levels and more frequently presented with splenomegaly. The clinical and biological features associated to progression-free and overall survival were: age, presence of splenomegaly, histotype (MF vs. PV vs. ET), anaemia, high platelet count and presence of any AD at diagnosis of MPN. The age-adjusted hazard ratio (HR) of progression for the presence of AD at diagnosis of MPN was 2.76. Overall survival was not significantly associated to AD at diagnosis, but the HR of progression for the presence of AD at diagnosis of MPN was 2.18. CONCLUSIONS: A possible common genetic predisposition, the inflammatory bone marrow microenvironment and the activation of theJAK/STAT pathway could be considered as responsible for the observed association between MPNs and ADs.

Development and application of a diagnostic and severity scale to grade post-operative pediatric cerebellar mutism syndrome.

The post-operative pediatric cerebellar mutism syndrome (CMS) affects about one-third of children and adolescents following surgical removal of a posterior fossa tumor (PFT). According to the Posterior Fossa Society consensus working definition, CMS is characterized by delayed-onset mutism/reduced speech and emotional lability after cerebellar or 4th ventricle tumor surgery in children, and is frequently accompanied by additional features such as hypotonia and oropharyngeal dysfunction/dysphagia. The main objective of this work was to develop a diagnostic scale to grade CMS duration and severity. Thirty consecutively referred subjects, aged 1-17 years (median 8 years, IQR 3-10), were evaluated with the proposed Post-Operative Pediatric CMS Survey after surgical resection of a PFT and, in case of CMS, for 30 days after the onset (T0) or until symptom remission. At day 30 (T1), CMS was classified into mild, moderate, or severe according to the proposed scale. CMS occurred in 13 patients (43%, 95% C.I.: 25.5-62.6%), with mild severity in 4 cases (31%), moderate in 4 (31%), and severe in 5 (38%). At T1, longer symptom persistence was associated with greater severity (p = 0.01). Greater severity at T0 predicted greater severity at T1 (p = 0.0001). Children with a midline tumor location and those aged under 5 years at diagnosis were at higher risk of CMS (p = 0.025 and p = 0.008, respectively). In conclusion, the proposed scale is a simple and applicable tool for estimating the severity of CMS at its onset, monitoring its course over time, and providing an early prognostic stratification to guide treatment decisions.

Pediatric diffuse midline glioma H3K27- altered: A complex clinical and biological landscape behind a neatly defined tumor type.

The 2021 World Health Organization Classification of Tumors of the Central Nervous System, Fifth Edition (WHO-CNS5), has strengthened the concept of tumor grade as a combination of histologic features and molecular alterations. The WHO-CNS5 tumor type "Diffuse midline glioma, H3K27-altered," classified within the family of "Pediatric-type diffuse high-grade gliomas," incarnates an ideally perfect integrated diagnosis in which location, histology, and genetics clearly define a specific tumor entity. It tries to evenly characterize a group of neoplasms that occur primarily in children and midline structures and that have a dismal prognosis. Such a well-defined pathological categorization has strongly influenced the pediatric oncology community, leading to the uniform treatment of most cases of H3K27-altered diffuse midline gliomas (DMG), based on the simplification that the mutation overrides the histological, radiological, and clinical characteristics of such tumors. Indeed, multiple studies have described pediatric H3K27-altered DMG as incurable tumors. However, in biology and clinical practice, exceptions are frequent and complexity is the rule. First of all, H3K27 mutations have also been found in non-diffuse gliomas. On the other hand, a minority of DMGs are H3K27 wild-type but have a similarly poor prognosis. Furthermore, adult-type tumors may rarely occur in children, and differences in prognosis have emerged between adult and pediatric H3K27-altered DMGs. As well, tumor location can determine differences in the outcome: patients with thalamic and spinal DMG have significantly better survival. Finally, other concomitant molecular alterations in H3K27 gliomas have been shown to influence prognosis. So, when such additional mutations are found, which one should we focus on in order to make the correct clinical decision? Our review of the current literature on pediatric diffuse midline H3K27-altered DMG tries to address such questions. Indeed, H3K27 status has become a fundamental supplement to the histological grading of pediatric gliomas; however, it might not be sufficient alone to exhaustively define the complex biological behavior of DMG in children and might not represent an indication for a unique treatment strategy across all patients, irrespective of age, additional molecular alterations, and tumor location.

Free Fatty Acids Signature in Human Intestinal Disorders: Significant Association between Butyric Acid and Celiac Disease.

Altered circulating levels of free fatty acids (FFAs), namely short chain fatty acids (SCFAs), medium chain fatty acids (MCFAs), and long chain fatty acids (LCFAs), are associated with metabolic, gastrointestinal, and malignant diseases. Hence, we compared the serum FFA profile of patients with celiac disease (CD), adenomatous polyposis (AP), and colorectal cancer (CRC) to healthy controls (HC). We enrolled 44 patients (19 CRC, 9 AP, 16 CD) and 16 HC. We performed a quantitative FFA evaluation with the gas chromatography-mass spectrometry method (GC-MS), and we performed Dirichlet-multinomial regression in order to highlight disease-specific FFA signature. HC showed a different composition of FFAs than CRC, AP, and CD patients. Furthermore, the partial least squares discriminant analysis (PLS-DA) confirmed perfect overlap between the CRC and AP patients and separation of HC from the diseased groups. The Dirichlet-multinomial regression identified only strong positive association between CD and butyric acid. Moreover, CD patients showed significant interactions with age, BMI, and gender. In addition, among patients with the same age and BMI, being male compared to being female implies a decrease of the CD effect on the (log) prevalence of butyric acid in FFA composition. Our data support GC-MS as a suitable method for the concurrent analysis of circulating SCFAs, MCFAs, and LCFAs in different gastrointestinal diseases. Furthermore, and notably, we suggest for the first time that butyric acid could represent a potential biomarker for CD screening.

Diving into Inflammation: A Pilot Study Exploring the Dynamics of the Immune-Microbiota Axis in Ileal Tissue Layers of Patients with Crohn's Disease.

BACKGROUND AND AIMS: The pathogenesis of Crohn's disease [CD] is still unclear. Disorders in the mucosal immunoregulation and its crosstalk with the microbiota may represent an important component in tissue injury. We aimed to characterize the molecular immune response distribution within the ileal layers and to evaluate the correlated microbiota in pathological/healthy settings comparing first surgery/relapse clinical conditions. METHODS: We enrolled 12 CD patients. A comprehensive analysis of an ileal mucosa, submucosa and serosa broad-spectrum cytokine panel was performed through a multiplex approach. In addition, ileal microbiota composition was assessed through next generation sequencing. RESULTS: We observed a distinct profile [of IL1-α, IL-1ß, IL-4, IL-8, ICAM-1, E-Selectin, P-Selectin, IP-10, IL 6 and IL 18] across the CD vs healthy ileal layers; and a different distribution of IFN- γ, P-Selectin, IL-27 and IL-21 in first surgery vs relapse patients. In addition, the phylum Tenericutes, the family Ruminococcaceae, and the genera Mesoplasma and Mycoplasma were significantly enriched in the pathological setting. Significant microbiota differences were observed between relapse and first surgery patients regarding the class Bacteroidia, and the genera Prevotella, Flavobacterium, Tepidimonas and Escherichia/Shigella. Finally, the abundance of the genus Mycoplasma was positively correlated with IL-18. CONCLUSIONS: We describe a dissimilarity of cytokine distribution and microbiota composition within CD and adjacent healthy ileal tissue layers and between first operation and surgical relapse. Our results give potential insight into the dynamics of the gut microbiota-immune axis in CD patients, leading to detection of new biomarkers.

Tyrosine Kinase Inhibitors Play an Antiviral Action in Patients Affected by Chronic Myeloid Leukemia: A Possible Model Supporting Their Use in the Fight Against SARS-CoV-2.

SARS-CoV-2 is the viral agent responsible for the pandemic that in the first months of 2020 caused about 400,000 deaths. Among compounds proposed to fight the SARS-CoV-2-related disease (COVID-19), tyrosine kinase inhibitors (TKIs), already effective in Philadelphia-positive acute lymphoblastic leukemia (Ph+ ALL) and chronic myeloid leukemia (CML), have been proposed on the basis of their antiviral action already demonstrated against SARS-CoV-1. Very few cases of COVID-19 have been reported in Ph+ ALL and in CML Italian cohorts; authors suggested that this low rate of infections might depend on the use of TKIs, but the biological causes of this phenomenon remain unknown. In this study, the CML model was used to test if TKIs would sustain or not the viral replication and if they could damage patient immunity. Firstly, the infection and replication rate of torquetenovirus (TTV), whose load is inversely proportional to the host immunological control, have been measured in CML patients receiving nilotinib. A very low percentage of subjects were infected at baseline, and TTV did not replicate or at least showed a low replication rate during the follow-up, with a mean load comparable to the measured one in healthy subjects. Then, after gene expression profiling experiments, we found that several "antiviral" genes, such as CD28 and IFN gamma, were upregulated, while genes with "proviral" action, such as ARG-1, CEACAM1, and FUT4, were less expressed during treatment with imatinib, thus demonstrating that TKIs are not detrimental from the immunological point of view. To sum up, our data could offer some biological explanations to the low COVID-19 occurrence in Ph+ ALL and CML patients and sustain the use of TKIs in COVID-19, as already proposed by several international ongoing studies.

The Italian consensus conference on the role of rehabilitation for children and adolescents with leukemia, central nervous system, and bone tumors, part 1: Review of the conference and presentation of consensus statements on rehabilitative evaluation of motor aspects.

Because of increasing survival rates in pediatric oncology, attention is focusing on cancer and its treatment-related side effects. Rehabilitation may reduce their impact. However, the literature does not provide strong evidence regarding rehabilitation pathways. Therefore, the Italian Association of Pediatric Hematology and Oncology organized a consensus conference on the role of rehabilitation of motor impairments in children/adolescents affected by leukemia, central nervous system, and bone tumors to define recommendations for daily practice. The grading of recommendation assessment, developing and evaluation (GRADE) method was used in order to formulate questions, select outcomes, evaluate evidence, and create recommendations. This paper includes the results on the rehabilitation assessment.

Organ Stiffness in the Work-Up of Myelofibrosis and Philadelphia-Negative Chronic Myeloproliferative Neoplasms.

To define the role of spleen stiffness (SS) and liver stiffness (LS) in myelofibrosis and other Philadelphia (Ph)-negative myeloproliferative neoplasms (MPNs), we studied, by ultrasonography (US) and elastography (ES), 70 consecutive patients with myelofibrosis (MF) (no.43), essential thrombocythemia (ET) (no.10), and polycythemia vera (PV) (no.17). Overall, the median SS was not different between patients with MF and PV (p = 0.9); however, both MF and PV groups had significantly higher SS than the ET group (p = 0.011 and p = 0.035, respectively) and healthy controls (p < 0.0001 and p = 0.002, respectively). In patients with MF, SS values above 40 kPa were significantly associated with worse progression-free survival (PFS) (p = 0.012; HR = 3.2). SS also correlated with the extension of bone marrow fibrosis (BMF) (p < 0.0001). SS was higher in advanced fibrotic stages MF-2, MF-3 (W.H.O. criteria) than in pre-fibrotic/early fibrotic stages (MF-0, MF-1) (p < 0.0001) and PFS was significantly different in the two cohorts, with values of 63% and 85%, respectively (p = 0.038; HR = 2.61). LS significantly differed between the patient cohort with MF and healthy controls (p = 0.001), but not between the patient cohorts with ET and PV and healthy controls (p = 0.999 and p = 0.101, respectively). We can conclude that organ stiffness adds valuable information to the clinical work-up of MPNs and could be employed to define patients at a higher risk of progression.

A Structurally Simple Vaccine Candidate Reduces Progression and Dissemination of Triple-Negative Breast Cancer.

The Tn antigen is a well-known tumor-associated carbohydrate determinant, often incorporated in glycopeptides to develop cancer vaccines. Herein, four copies of a conformationally constrained mimetic of the antigen TnThr (GalNAc-Thr) were conjugated to the adjuvant CRM197, a protein licensed for human use. The resulting vaccine candidate, mime[4]CRM elicited a robust immune response in a triple-negative breast cancer mouse model, correlated with high frequency of CD4+ T cells and low frequency of M2-type macrophages, which reduces tumor progression and lung metastasis growth. Mime[4]CRM-mediated activation of human dendritic cells is reported, and the proliferation of mime[4]CRM-specific T cells, in cancer tissue and peripheral blood of patients with breast cancer, is demonstrated. The locked conformation of the TnThr mimetic and a proper presentation on the surface of CRM197 may explain the binding of the conjugate to the anti-Tn antibody Tn218 and its efficacy to fight cancer cells in mice.

Erythropoietin treatment in chronic phase chronic myeloid leukemia patients treated with frontline imatinib who developed late anemia.

BACKGROUND: Role of erythropoietin (EPO) in the treatment of late anemia in patients with Chronic Myeloid Leukemia (CML) is still undefined. METHODS: Fifty CML patients treated at 14 institutions with frontline imatinib for at least 12 months and in stable complete cytogenetic response who developed a late chronic anemia treated with EPO were retrospectively evaluated. RESULTS: Median time from imatinib start to EPO treatment was 42.2 months [interquartile range (IQR) 20.8-91.9]. Median Hb value at EPO starting time was 9.9 g/dL (IQR 8.9-10.3): Eleven patients (22.0%) were transfusion dependent. Alpha-EPO (40 000 UI weekly) was employed in 37 patients, beta-EPO (30 000 UI weekly) in 9 patients, zeta-EPO (40 000 UI weekly) in 2 patients, and darbepoetin (150 mcg/weekly) in the remaining 2 patients. On the whole, 41 patients (82.0%) achieved an erythroid response, defined as a stable (>3 months) improvement >1.5 g/dL of Hb level, and 9 patients (18.0%) indeed resulted resistant. Among responding patients, 10 relapsed after a median time from EPO start of 20.7 months (IQR 10.8-63.7). No EPO-related toxicity was observed. CONCLUSIONS: Results of EPO treatment for late chronic anemia during long-lasting imatinib therapy are encouraging, with a high rate of response.

Significant and Conflicting Correlation of IL-9 With Prevotella and Bacteroides in Human Colorectal Cancer.

Background and aim: Gut microbiota (GM) can support colorectal cancer (CRC) progression by modulating immune responses through the production of both immunostimulatory and/or immunosuppressive cytokines. The role of IL-9 is paradigmatic because it can either promote tumor progression in hematological malignancies or inhibit tumorigenesis in solid cancers. Therefore, we investigate the microbiota-immunity axis in healthy and tumor mucosa, focusing on the correlation between cytokine profile and GM signature. Methods: In this observational study, we collected tumor (CRC) and healthy (CRC-S) mucosa samples from 45 CRC patients, who were undergoing surgery in 2018 at the Careggi University Hospital (Florence, Italy). First, we characterized the tissue infiltrating lymphocyte subset profile and the GM composition. Subsequently, we evaluated the CRC and CRC-S molecular inflammatory response and correlated this profile with GM composition, using Dirichlet multinomial regression. Results: CRC samples displayed higher percentages of Th17, Th2, and Tregs. Moreover, CRC tissues showed significantly higher levels of MIP-1α, IL-1α, IL-1ß, IL-2, IP-10, IL-6, IL-8, IL-17A, IFN-γ, TNF-α, MCP-1, P-selectin, and IL-9. Compared to CRC-S, CRC samples also showed significantly higher levels of the following genera: Fusobacteria, Proteobacteria, Fusobacterium, Ruminococcus2, and Ruminococcus. Finally, the abundance of Prevotella spp. in CRC samples negatively correlated with IL-17A and positively with IL-9. On the contrary, Bacteroides spp. presence negatively correlated with IL-9. Conclusions: Our data consolidate antitumor immunity impairment and the presence of a distinct microbiota profile in the tumor microenvironment compared with the healthy mucosa counterpart. Relating the CRC cytokine profile with GM composition, we confirm the presence of bidirectional crosstalk between the immune response and the host's commensal microorganisms. Indeed, we document, for the first time, that Prevotella spp. and Bacteroides spp. are, respectively, positively and negatively correlated with IL-9, whose role in CRC development is still under debate.

Evaluation and comparison of short chain fatty acids composition in gut diseases.

BACKGROUND: An altered (dysbiosis) and unhealthy status of the gut microbiota is usually responsible for a reduction of short chain fatty acids (SCFAs) concentration. SCFAs obtained from the carbohydrate fermentation processes are crucial in maintaining gut homeostasis and their determination in stool samples could provide a faster, reliable and cheaper method to highlight the presence of an intestinal dysbiosis and a biomarker for various gut diseases. We hypothesize that different intestinal diseases, such as celiac disease (CD), adenomatous polyposis (AP) and colorectal cancer (CRC) could display a particular fecal SCFAs' signature. AIM: To compare the fecal SCFAs' profiles of CD, AP, CRC patients and healthy controls, using the same analytical method. METHODS: In this cross-sectional study, we defined and compared the SCFAs' concentration in fecal samples of 9 AP, 16 CD, 19 CRC patients and 16 healthy controls (HC). The SCFAs' analysis were performed using a gas-chromatography coupled with mass spectrometry method. Data analysis was carried out using Wilcoxon rank-sum test to assess pairwise differences of SCFAs' profiles, partial least squares-discriminate analysis (PLS-DA) to determine the status membership based on distinct SCFAs' profiles, and Dirichlet regression to determine factors influencing concentration levels of SCFAs. RESULTS: We have not observed any difference in the SCFAs' amount and composition between CD and healthy control. On the contrary, the total amount of SCFAs was significantly lower in CRC patients compared to HC (P = 0.044) and CD (P = 0.005). Moreover, the SCFAs' percentage composition was different in CRC and AP compared to HC. In detail, HC displayed higher percentage of acetic acid (P value = 1.3 × 10-6) and a lower amount of butyric (P value = 0.02192), isobutyric (P value = 7.4 × 10-5), isovaleric (P value = 0.00012) and valeric (P value = 0.00014) acids compared to CRC patients. AP showed a lower abundance of acetic acid (P value = 0.00062) and higher percentages of propionic (P value = 0.00433) and isovaleric (P value = 0.00433) acids compared to HC. Moreover, AP showed higher levels of propionic acid (P value = 0.03251) and a lower level of isobutyric acid (P value = 0.00427) in comparison to CRC. The PLS-DA model demonstrated a significant separation of CRC and AP groups from HC, although some degree of overlap was observed between CRC and AP. CONCLUSION: Analysis of fecal SCFAs shows the potential to provide a non-invasive means of diagnosis to detect patients with CRC and AP, while CD patients cannot be discriminated from healthy subjects.