Evaluation d'une méthode de diagnostic des anomalies de la bêta-oxydation mitochondriale sur fibroblastes par spectrométrie de masse de type Electrospray-MS/M

LILLE2-BU Santé-Recherche (593502101) / SudocSudocFranceF

Induction par le couple MIF-CXCR4 d'un phénotype invasif et métastatique au sein de cellules tumorales coliques humaines chimiorésistantes

Contexte : Malgré le développement de thérapies combinées utilisant des agents anticancéreux cytotoxiques et/ou des thérapies ciblant les voies de signalisation oncogénique, la persistance de cellules tumorales agressives reste un obstacle majeur au traitement des cancers. Objectif : Le but de ce travail était d'analyser les mécanismes moléculaires de la survie et de l'agressivité métastatique de cellules cancéreuses coliques chimiorésistantes. Méthode : La recherche d éle ments impliqués dans l agressivité métastatique et la résistance aux drogues a été réalisée en combinant les approches de transcriptome, RT-PCR quantitative, Western Blot, ELISA et cytométrie en flux. L étude du potentiel métastatique a été effectuée en utilisant les techniques d invasion in vitro sur collagène de type I et de xénogreffes sous-cutanées sur modèles murins. L étude des mécanismes responsables a fait appel à des inhibiteurs pharmacologiques, des anticorps bloquants et à la technique d invalidation génique par ARN interférence. Résultats : Nous avons caractérisé des cellules HT-29 chimiorésistantes spontanément invasives dans une matrice de collagène de type I et spontanément métastatiques in vivo dans des modèles de xénogreffes souscutanées, versus des cellules non invasives et non métastatiques. Une approche transcriptomique comparative sur puces pangénomiques a permis de mettre en évidence la surexpression très importante d un récepteur de chimiokine, CXCR4, dans les cellules invasives comparativement aux cellules non invasives. La cytokine MIF a été identifiée comme le ligand autocrine de CXCR4 responsable de l invasion cellulaire. L invalidation de CXCR4 et le ciblage pharmacologique de l axe MIF/CXCR4 abolissent ce phénotype agressif. L induction de CXCR4 est associée à la surexpression de HIF-2! et ASCL2, deux facteurs de transcription impliqués dans le contrôle de l expression de CXCR4 et la maintenance des cellules souches intestinales. Une étude pilote effectuée sur une série de métastases hépatiques issues de patients atteints de cancer colorectal a révélé une augmentation significative et importante des transcrits de CXCR4 chez les patients ayant suivi une chimiothérapie néo-adjuvante par le protocole FOLFOX. Conclusion : Le ciblage de l axe CXCR4/MIF en association thérapeutique pour le traitement du cancer colorectal pourrait contrecarrer l émergence d un comportement invasif et métastatique au sein de populations clones de cellules tumorales coliques chimiorésistantes.LILLE2-BU Santé-Recherche (593502101) / SudocSudocFranceF

Pregnancy in metabolic diseases with hepatic expression what risks for the mother and the child?

International audienceInborn errors of metabolism (IEM) are rare diseases caused by mutations in genes encoding enzymes or carriers. Qualitative or quantitative protein deficiency induces both an accumulation of precursor metabolites and a lack of products downstream of the blockade. Pregnancy in patients with IEM is a condition likely to promote metabolic decompensation. In this review, we presented liver symptoms described during pregnancy in a context of hepatic IEM. In particular, we detailed clinical and biological abnormalities specifically occurring in tyrosinemia type I, Wilson disease, and main urea cycle defects. In the case of hepatic IEM, depending on the deficit, pregnant women have an increased risk of pre-eclampsia and HELLP syndrome, as well as hyperammonemia. Wilson disease, and principal urea cycle defects. Multidisciplinary consultation is essential for the optimal management of pregnant women with IEM as well as newborns

First-line Screening of OXPHOS Deficiencies Using Microscale Oxygraphy in Human Skin Fibroblasts: A Preliminary Study

International audienceThe diagnosis of mitochondrial diseases is a real challenge because of the vast clinical and genetic heterogeneity. Classically, the clinical examination and genetic analysis must be completed by several biochemical assays to confirm the diagnosis of mitochondrial disease. Here, we tested the validity of microscale XF technology in measuring oxygen consumption in human skin fibroblasts isolated from 5 pediatric patients with heterogeneous mitochondrial disorders. We first set up the protocol conditions to allow the determination of respiratory parameters including respiration associated with ATP production, proton leak, maximal respiration, and spare respiratory capacity with reproducibility and repeatability. Maximum respiration and spare capacity were the only parameters decreased in patients irrespective of the type of OXPHOS deficiency. These results were confirmed by high-resolution oxygraphy, the reference method to measure cellular respiration. Given the fact that microscale XF technology allows fast, automated and standardized measurements, we propose to use microscale oxygraphy among the first-line methods to screen OXPHOS deficiencies

Long-term prognosis of fatty-acid oxidation disorders in adults: Optimism despite the limited effective therapies available

International audienceIntroduction: Fatty-acid oxidation disorders (FAODs) are recessive genetic diseases. Materials and methods: We report here clinical and paraclinical data from a retrospective study of 44 adults with muscular FAODs from six French reference centers for neuromuscular or metabolic diseases. Results: The study cohort consisted of 44 adult patients: 14 with carnitine palmitoyl transferase 2 deficiency (32%), nine with multiple acyl-CoA deficiency (20%), 13 with very long-chain acyl-CoA dehydrogenase deficiency (30%), three with long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (7%), and five with short-chain acyl-CoA dehydrogenase deficiency (11%). Disease onset occurred during childhood in the majority of patients (59%), with a mean age at onset of 15 years (range = 0.5–35) and a mean of 12.6 years (range = 0–58) from disease onset to diagnosis. The principal symptoms were acute muscle manifestations (rhabdomyolysis, exercise intolerance, myalgia), sometimes associated with permanent muscle weakness. Episodes of rhabdomyolysis were frequent (84%), with a mean creatinine kinase level of 68,958 U/L (range = 660–300,000). General metabolic complications were observed in 58% of patients, respiratory manifestations in 18% of cases, and cardiological manifestations in 9% of cases. Fasting acylcarnitine profile was used to orient genetic explorations in 65% of cases. After a mean follow-up of 10 years, 33% of patients were asymptomatic and 56% continued to display symptoms after exercise. The frequency of rhabdomyolysis decreased after diagnosis in 64% of cases. Conclusion: A standardized register would complete this cohort description of muscular forms of FAODs with exhaustive data, making it possible to assess the efficacy of therapeutic protocols in real-life conditions and during the long-term follow-up of patients

Newborn Screening of Primary Carnitine Deficiency: An Overview of Worldwide Practices and Pitfalls to Define an Algorithm before Expansion of Newborn Screening in France

Primary Carnitine Deficiency (PCD) is a fatty acid oxidation disorder that will be included in the expansion of the French newborn screening (NBS) program at the beginning of 2023. This disease is of high complexity to screen, due to its pathophysiology and wide clinical spectrum. To date, few countries screen newborns for PCD and struggle with high false positive rates. Some have even removed PCD from their screening programs. To understand the risks and pitfalls of implementing PCD to the newborn screening program, we reviewed and analyzed the literature to identify hurdles and benefits from the experiences of countries already screening this inborn error of metabolism. In this study, we therefore, present the main pitfalls encountered and a worldwide overview of current practices in PCD newborn screening. In addition, we address the optimized screening algorithm that has been determined in France for the implementation of this new condition

Autocrine Induction of Invasive and Metastatic Phenotypes by the MIF-CXCR4 Axis in Drug-Resistant Human Colon Cancer Cells

International audienceMetastasis and drug resistance are major problems in cancer chemotherapy. The purpose of this work was to analyze the molecular mechanisms underlying the invasive potential of drug-resistant colon carcinoma cells. Cellular models included the parental HT-29 cell line and its drug-resistant derivatives selected after chronic treatment with either 5-fluorouracil, methotrexate, doxorubicin, or oxaliplatin. Drug-resistant invasive cells were compared with noninvasive cells using cDNA microarray, quantitative reverse transcription-PCR, flow cytometry, immunoblots, and ELISA. Functional and cellular signaling analyses were undertaken using pharmacologic inhibitors, function-blocking antibodies, and silencing by retrovirus-mediated RNA interference. 5-Fluorouracil- and methotrexate-resistant HT-29 cells expressing an invasive phenotype in collagen type I and a metastatic behavior in immunodeficient mice exhibited high expression of the chemokine receptor CXCR4. Macrophage migration-inhibitory factor (MIF) was identified as the critical autocrine CXCR4 ligand promoting invasion in drug-resistant colon carcinoma HT-29 cells. Silencing of CXCR4 and impairing the MIF-CXCR4 signaling pathways by ISO-1, pAb FL-115, AMD-3100, monoclonal antibody 12G5, and BIM-46187 abolished this aggressive phenotype. Induction of CXCR4 was associated with the upregulation of two genes encoding transcription factors previously shown to control CXCR4 expression (HIF-2alpha and ASCL2) and maintenance of intestinal stem cells (ASCL2). Enhanced CXCR4 expression was detected in liver metastases resected from patients with colon cancer treated by the standard FOLFOX regimen. Combination therapies targeting the CXCR4-MIF axis could potentially counteract the emergence of the invasive metastatic behavior in clonal derivatives of drug-resistant colon cancer cells

Omics to Explore Amyotrophic Lateral Sclerosis Evolution: the Central Role of Arginine and Proline Metabolism

International audienc