[Artificial intelligence: to a better predictive strategy for testicular sperm extraction outcome in azoospermia]. / Intelligence artificielle : vers une meilleure stratégie de prédiction du résultat de la biopsie testiculaire dans un contexte d'azoospermie.

Azoospermia, defined as the absence of sperm in the semen, is found in 10-15 % of infertile patients. Two-thirds of these cases are caused by impaired spermatogenesis, known as non-obstructive azoospermia (NOA). In this context, surgical sperm extraction using testicular sperm extraction (TESE) is the best option and can be offered to patients as part of fertility preservation, or to benefit from in vitro fertilization. The aim of the preoperative assessment is to identify the cause of NOA and evaluate the status of spermatogenesis. Its capacity to predict TESE success remains limited. As a result, no objective and reliable criteria are currently available to guide professionals on the chances of success and enable them to correctly assess the benefit-risk balance of this procedure. Artificial intelligence (AI), a field of research that has been rapidly expanding in recent years, has the potential to revolutionize medicine by making it more predictive and personalized. The aim of this review is to introduce AI and its key concepts, and then to examine the current state of research into predicting the success of TESE.

Machine learning approach to assess the association between anthropometric, metabolic, and nutritional status and semen parameters.

ABSTRACT: Many lifestyle factors, such as nutritional imbalance leading to obesity, metabolic disorders, and nutritional deficiency, have been identified as potential risk factors for male infertility. The aim of this study was to evaluate the relationship between semen parameters and anthropometric, metabolic and nutritional parameters. Relationship was first assessed individually, then after the application of a previously constructed and validated machine learning score that allows their combination. Anthropometric, metabolic, antioxidant, micronutrient, and sperm parameters from 75 men suffering from idiopathic infertility from four infertility centers in France (Jean-Verdier ART Center Hospital, Bondy; North Hospital ART Center, Saint-Étienne; Navarre Polyclinic ART Center, Pau; and Cochin Hospital ART Center, Paris) between September 2009 and December 2013 were collected. After assessing standard correlation analysis, a previously built machine learning model, providing a score ranging from 0 (the poorest) to 1 (the most favorable), was calculated for each man in the study cohort. This machine learning model, which separates infertile/fertile men with unexplained infertility on the basis of their bioclinical signature, provides a more holistic evaluation of the influence of the considered markers (anthropometric, metabolic, and oxidative status). We observed a significant correlation of some anthropometric, metabolic, and nutritional disorders with some sperm characteristics. Moreover, an unfavorable machine learning score was associated with a high level of sperm DNA fragmentation. Favorable anthropometric, metabolic, and oxidative patterns, which may reflect an appropriate lifestyle, appear to positively impact overall health, in particular reproductive function. This study, consistent with previous publications, suggests that beyond semen quality parameters, in an essential assessment of male fertility, other key factors should be taken into account. In this regard, the application of emerging artificial intelligence techniques may provide a unique opportunity to integrate all these parameters and deliver personalized care.

Dose-response evaluation of intravenous gene therapy in a symptomatic mouse model of metachromatic leukodystrophy.

Metachromatic leukodystrophy (MLD) is a rare, autosomal recessive neurodegenerative disease caused by deficient activity of the lysosomal enzyme arylsulfatase A (ARSA), resulting in sulfatide accumulation and subsequent demyelination and neuronal damage within the central and peripheral nervous systems. Three clinical forms of MLD have been described, based on age at symptom onset. The most frequent and severe forms have an early onset, with the disease progressing rapidly toward severe motor and cognitive regression and ultimately premature death. There are currently no approved therapies for most of these early-onset patients once symptoms are present. Thus, it is crucial to develop new approaches to treat symptomatic patients. Here, we proposed a gene therapy approach based on the intravenous delivery of AAVPHP.eB encoding ARSA. MLD mice were treated at 6 months for a dose-response study and at 9 months to assess late-treatment efficacy. Therapeutic efficacy was evaluated 3 or 6 months after injection. We demonstrated a broad transduction in the central nervous system, a complete correction of sulfatide storage, and a significant improvement in neuroinflammation at low dose and late treatment. Taken together, this work establishes a strong rationale for proposing a phase I/II clinical trial in MLD patients.

Effect of Stool Sampling on a Routine Clinical Method for the Quantification of Six Short Chain Fatty Acids in Stool Using Gas Chromatography-Mass Spectrometry.

Short chain fatty acids (SCFAs) are primarily produced in the caecum and proximal colon via the bacterial fermentation of undigested carbohydrates that have avoided digestion in the small intestine. Increasing evidence supports the critical role that SCFAs play in health and homeostasis. Microbial SCFAs, namely butyric acid, serve as a principal energy source for colonocytes, and their production is essential for gut integrity. A direct link between SCFAs and some human pathological conditions, such as inflammatory bowel disease, irritable bowel syndrome, diarrhea, and cancer, has been proposed. The direct measurement of SCFAs in feces provides a non-invasive approach to demonstrating connections between SCFAs, microbiota, and metabolic diseases to estimate their potential applicability as meaningful biomarkers of intestinal health. This study aimed to adapt a robust analytical method (liquid-liquid extraction, followed by isobutyl chloroformate derivatization and GC-MS analysis), with comparable performances to methods from the literature, and to use this tool to tackle the question of pre-analytical conditions, namely stool processing. We focused on the methodology of managing stool samples before the analysis (fresh stool or dilution in either ethanol/methanol, lyophilized stool, or RNAlater®), as this is a significant issue to consider for standardizing results between clinical laboratories. The objective was to standardize methods for future applications as diagnostic tools. In this paper, we propose a validated GC-MS method for SCFA quantification in stool samples, including pre- and post-analytical comparison studies that could be easily used for clinical laboratory purposes. Our results show that using lyophilization as a stool-processing method would be the best method to achieve this goal.

Perturbation and resilience of the gut microbiome up to 3 months after ß-lactams exposure in healthy volunteers suggest an important role of microbial ß-lactamases.

BACKGROUND: Antibiotics notoriously perturb the gut microbiota. We treated healthy volunteers either with cefotaxime or ceftriaxone for 3 days, and collected in each subject 12 faecal samples up to day 90. Using untargeted and targeted phenotypic and genotypic approaches, we studied the changes in the bacterial, phage and fungal components of the microbiota as well as the metabolome and the ß-lactamase activity of the stools. This allowed assessing their degrees of perturbation and resilience. RESULTS: While only two subjects had detectable concentrations of antibiotics in their faeces, suggesting important antibiotic degradation in the gut, the intravenous treatment perturbed very significantly the bacterial and phage microbiota, as well as the composition of the metabolome. In contrast, treatment impact was relatively low on the fungal microbiota. At the end of the surveillance period, we found evidence of resilience across the gut system since most components returned to a state like the initial one, even if the structure of the bacterial microbiota changed and the dynamics of the different components over time were rarely correlated. The observed richness of the antibiotic resistance genes repertoire was significantly reduced up to day 30, while a significant increase in the relative abundance of ß-lactamase encoding genes was observed up to day 10, consistent with a concomitant increase in the ß-lactamase activity of the microbiota. The level of ß-lactamase activity at baseline was positively associated with the resilience of the metabolome content of the stools. CONCLUSIONS: In healthy adults, antibiotics perturb many components of the microbiota, which return close to the baseline state within 30 days. These data suggest an important role of endogenous ß-lactamase-producing anaerobes in protecting the functions of the microbiota by de-activating the antibiotics reaching the colon. Video Abstract.

Successful treatment of severe MSUD in Bckdhb-/- mice with neonatal AAV gene therapy.

Maple syrup urine disease (MSUD) is rare autosomal recessive metabolic disorder caused by the dysfunction of the mitochondrial branched-chain 2-ketoacid dehydrogenase (BCKD) enzyme complex leading to massive accumulation of branched-chain amino acids and 2-keto acids. MSUD management, based on a life-long strict protein restriction with nontoxic amino acids oral supplementation represents an unmet need as it is associated with a poor quality of life, and does not fully protect from acute life-threatening decompensations or long-term neuropsychiatric complications. Orthotopic liver transplantation is a beneficial therapeutic option, which shows that restoration of only a fraction of whole-body BCKD enzyme activity is therapeutic. MSUD is thus an ideal target for gene therapy. We and others have tested AAV gene therapy in mice for two of the three genes involved in MSUD, BCKDHA and DBT. In this study, we developed a similar approach for the third MSUD gene, BCKDHB. We performed the first characterization of a Bckdhb-/- mouse model, which recapitulates the severe human phenotype of MSUD with early-neonatal symptoms leading to death during the first week of life with massive accumulation of MSUD biomarkers. Based on our previous experience in Bckdha-/- mice, we designed a transgene carrying the human BCKDHB gene under the control of a ubiquitous EF1α promoter, encapsidated in an AAV8 capsid. Injection in neonatal Bckdhb-/- mice at 1014 vg/kg achieved long-term rescue of the severe MSUD phenotype of Bckdhb-/- mice. These data further validate the efficacy of gene therapy for MSUD opening perspectives towards clinical translation.

Prospective comparison of diagnostic tests for bile acid diarrhoea.

BACKGROUND: Bile acid diarrhoea is often missed because gold standard nuclear medicine tauroselcholic [75-Se] acid (SeHCAT) testing has limited availability. Empirical treatment effect has unknown diagnostic performance, whereas plasma 7α-hydroxy-4-cholesten-3-one (C4) is inexpensive but lacks sensitivity. AIMS: To determine diagnostic characteristics of empirical treatment and explore improvements in diagnostics with potential better availability than SeHCAT. METHODS: This diagnostic accuracy study was part of a randomised, placebo-controlled trial of colesevelam. Consecutive patients with chronic diarrhoea attending SeHCAT had blood and stool sampled. Key thresholds were C4 > 46 ng/mL and SeHCAT retention ≤10%. A questionnaire recorded patient-reported empirical treatment effect. We analysed receiver operating characteristics and explored machine learning applied logistic regression and decision tree modelling with internal validation. RESULTS: Ninety-six (38%) of 251 patients had SeHCAT retention ≤10%. The effect of empirical treatment assessed with test results for bile acid studies blinded had 63% (95% confidence interval 44%-79%) sensitivity and 65% (47%-80%) specificity; C4 > 46 ng/mL had 47% (37%-57%) and 92% (87%-96%), respectively. A decision tree combining C4 ≥ 31 ng/mL with ≥1.1 daily watery stools (Bristol type 6 and 7) had 70% (51%-85%) sensitivity and 95% (83%-99%) specificity. The logistic regression model, including C4, the sum of measured stool bile acids and daily watery stools, had 77% (58%-90%) sensitivity and 93% (80%-98%) specificity. CONCLUSIONS: Diagnosis of bile acid diarrhoea using empirical treatment was inadequate. Exploration suggested considerable improvements in the sensitivity of C4-based testing, offering potential widely available diagnostics. Further validation is warranted. CLINICALTRIALS: gov: NCT03876717.

Membrane Lipids in Ultra-High-Risk Patients: Potential Predictive Biomarkers of Conversion to Psychosis.

Alterations in membrane lipids are reported in schizophrenia. However, no conclusion can be drawn regarding the extended and predictive value of these alterations in persons at ultra-high risk of psychosis (UHR). Recent studies suggested that sterols' impact on psychiatric disorders was underestimated. Here, we simultaneously explored sterols, fatty acids (FA), and phospholipids (PL) in UHR persons for the first time. We analysed erythrocyte membrane lipids in 61 UHR persons, including 29 who later converted to psychosis (UHR-C) and 32 who did not (UHC-NC). We used gas chromatography for FA and liquid chromatography tandem with mass spectrometry for sterols and phospholipids. Among UHR individuals, elevated baseline membrane linoleic acid level was associated with conversion to psychosis (26.1% vs. 60.5%, p = 0.02). Combining sterols, FA, and PL membrane composition improved the prediction of psychosis onset (AUC = 0.73). This is the first report showing that membrane sterol participates, with other membrane lipids, in modulating the risk of psychosis. It suggests that membrane lipids could be used as biomarkers for personalised medicine in UHR patients.

A Machine Learning Approach for the Prediction of Testicular Sperm Extraction in Nonobstructive Azoospermia: Algorithm Development and Validation Study.

BACKGROUND: Testicular sperm extraction (TESE) is an essential therapeutic tool for the management of male infertility. However, it is an invasive procedure with a success rate up to 50%. To date, no model based on clinical and laboratory parameters is sufficiently powerful to accurately predict the success of sperm retrieval in TESE. OBJECTIVE: The aim of this study is to compare a wide range of predictive models under similar conditions for TESE outcomes in patients with nonobstructive azoospermia (NOA) to identify the correct mathematical approach to apply, most appropriate study size, and relevance of the input biomarkers. METHODS: We analyzed 201 patients who underwent TESE at Tenon Hospital (Assistance Publique-Hôpitaux de Paris, Sorbonne University, Paris), distributed in a retrospective training cohort of 175 patients (January 2012 to April 2021) and a prospective testing cohort (May 2021 to December 2021) of 26 patients. Preoperative data (according to the French standard exploration of male infertility, 16 variables) including urogenital history, hormonal data, genetic data, and TESE outcomes (representing the target variable) were collected. A TESE was considered positive if we obtained sufficient spermatozoa for intracytoplasmic sperm injection. After preprocessing the raw data, 8 machine learning (ML) models were trained and optimized on the retrospective training cohort data set: The hyperparameter tuning was performed by random search. Finally, the prospective testing cohort data set was used for the model evaluation. The metrics used to evaluate and compare the models were the following: sensitivity, specificity, area under the receiver operating characteristic curve (AUC-ROC), and accuracy. The importance of each variable in the model was assessed using the permutation feature importance technique, and the optimal number of patients to include in the study was assessed using the learning curve. RESULTS: The ensemble models, based on decision trees, showed the best performance, especially the random forest model, which yielded the following results: AUC=0.90, sensitivity=100%, and specificity=69.2%. Furthermore, a study size of 120 patients seemed sufficient to properly exploit the preoperative data in the modeling process, since increasing the number of patients beyond 120 during model training did not bring any performance improvement. Furthermore, inhibin B and a history of varicoceles exhibited the highest predictive capacity. CONCLUSIONS: An ML algorithm based on an appropriate approach can predict successful sperm retrieval in men with NOA undergoing TESE, with promising performance. However, although this study is consistent with the first step of this process, a subsequent formal prospective multicentric validation study should be undertaken before any clinical applications. As future work, we consider the use of recent and clinically relevant data sets (including seminal plasma biomarkers, especially noncoding RNAs, as markers of residual spermatogenesis in NOA patients) to improve our results even more.

Roux-en-Y gastric bypass induces hepatic transcriptomic signatures and plasma metabolite changes indicative of improved cholesterol homeostasis.

BACKGROUND & AIMS: Roux-en-Y gastric bypass (RYGB), the most effective surgical procedure for weight loss, decreases obesity and ameliorates comorbidities, such as non-alcoholic fatty liver (NAFLD) and cardiovascular (CVD) diseases. Cholesterol is a major CVD risk factor and modulator of NAFLD development, and the liver tightly controls its metabolism. How RYGB surgery modulates systemic and hepatic cholesterol metabolism is still unclear. METHODS: We studied the hepatic transcriptome of 26 patients with obesity but not diabetes before and 1 year after undergoing RYGB. In parallel, we measured quantitative changes in plasma cholesterol metabolites and bile acids (BAs). RESULTS: RYGB surgery improved systemic cholesterol metabolism and increased plasma total and primary BA levels. Transcriptomic analysis revealed specific alterations in the liver after RYGB, with the downregulation of a module of genes implicated in inflammation and the upregulation of three modules, one associated with BA metabolism. A dedicated analysis of hepatic genes related to cholesterol homeostasis pointed towards increased biliary cholesterol elimination after RYGB, associated with enhancement of the alternate, but not the classical, BA synthesis pathway. In parallel, alterations in the expression of genes involved in cholesterol uptake and intracellular trafficking indicate improved hepatic free cholesterol handling. Finally, RYGB decreased plasma markers of cholesterol synthesis, which correlated with an improvement in liver disease status after surgery. CONCLUSIONS: Our results identify specific regulatory effects of RYGB on inflammation and cholesterol metabolism. RYGB alters the hepatic transcriptome signature, likely improving liver cholesterol homeostasis. These gene regulatory effects are reflected by systemic post-surgery changes of cholesterol-related metabolites, corroborating the beneficial effects of RYGB on both hepatic and systemic cholesterol homeostasis. IMPACT AND IMPLICATIONS: Roux-en-Y gastric bypass (RYGB) is a widely used bariatric surgery procedure with proven efficacy in body weight management, combatting cardiovascular disease (CVD) and non-alcoholic fatty liver disease (NAFLD). RYGB exerts many beneficial metabolic effects, by lowering plasma cholesterol and improving atherogenic dyslipidemia. Using a cohort of patients undergoing RYGB, studied before and 1 year after surgery, we analyzed how RYGB modulates hepatic and systemic cholesterol and bile acid metabolism. The results of our study provide important insights on the regulation of cholesterol homeostasis after RYGB and open avenues that could guide future monitoring and treatment strategies targeting CVD and NAFLD in obesity.

Enasidenib treatment in two individuals with D-2-hydroxyglutaric aciduria carrying a germline IDH2 mutation.

D-2-hydroxyglutaric aciduria type II (D2HGA2) is a severe inborn disorder of metabolism caused by heterozygous R140 mutations in the IDH2 (isocitrate dehydrogenase 2) gene. Here we report the results of treatment of two children with D2HGA2, one of whom exhibited severe dilated cardiomyopathy, with the selective mutant IDH2 enzyme inhibitor enasidenib. In both children, enasidenib treatment led to normalization of D-2-hydroxyglutarate (D-2-HG) concentrations in body fluids. At doses of 50 mg and 60 mg per day, no side effects were observed, except for asymptomatic hyperbilirubinemia. For the child with cardiomyopathy, chronic D-2-HG inhibition was associated with improved cardiac function, and for both children, therapy was associated with improved daily functioning, global motility and social interactions. Treatment of the child with cardiomyopathy led to therapy-coordinated changes in serum phospholipid levels, which were partly recapitulated in cultured fibroblasts, associated with complex effects on lipid and redox-related gene pathways. These findings indicate that targeted inhibition of a mutant enzyme can partly reverse the pathology of a chronic neurometabolic genetic disorder.

Adipocytes secretome from normal and tumor breast favor breast cancer invasion by metabolic reprogramming

Background Adipose tissue is a major component of breast stroma. This study focused on delineating the effects of adipose stem cells (ASCs) derived from breast of healthy women and cancer patients with normal or tumor breast cells. Methods The ASCs were induced to differentiate into adipocytes, and the subsequent adipocyte conditioned media (ACM) were evaluated for their fatty acid profile, adipokine secretion and influence on proliferation, migration and invasion on tumoral (MCF-7 and SUM159) and normal (HMEC) human breast cell lines. Results An enrichment of arachidonic acid was observed in ACM from tumor tissues. Adipose tissues from tumor free secrete twice as much leptin than those from proximal or distal to the tumor. All ACMs display proliferative activity and favor invasiveness of SUM159 cells compared to MCF-7 and HMEC. All ACMs induced lipid droplets accumulation in MCF-7 cells and increased CD36 expression in tumor cells. Conclusion We conclude that among secreted factors analyzed, only arachidonic acid and leptin levels did discriminate ASCs from tumor-bearing and tumor-free breasts emphasizing the importance that other cell types could contribute to the adipose tissue secretome in a tumor context (AU)

Comparing Transgenic Production to Supplementation of ω-3 PUFA Reveals Distinct But Overlapping Mechanisms Underlying Protection Against Metabolic and Hepatic Disorders.

We compared endogenous ω-3 PUFA production to supplementation for improving obesity-related metabolic dysfunction. Fat-1 transgenic mice, who endogenously convert exogenous ω-6 to ω-3 PUFA, and wild-type littermates were fed a high-fat diet and a daily dose of either ω-3 or ω-6 PUFA-rich oil for 12 wk. The endogenous ω-3 PUFA production improved glucose intolerance and insulin resistance but not hepatic steatosis. Conversely, ω-3 PUFA supplementation fully prevented hepatic steatosis but failed to improve insulin resistance. Both models increased hepatic levels of ω-3 PUFA-containing 2-monoacylglycerol and N-acylethanolamine congeners, and reduced levels of ω-6 PUFA-derived endocannabinoids with ω-3 PUFA supplementation being more efficacious. Reduced hepatic lipid accumulation associated with the endocannabinoidome metabolites EPEA and DHEA, which was causally demonstrated by lower lipid accumulation in oleic acid-treated hepatic cells treated with these metabolites. While both models induced a significant fecal enrichment of the beneficial Allobaculum genus, mice supplemented with ω-3 PUFA displayed additional changes in the gut microbiota functions with a significant reduction of fecal levels of the proinflammatory molecules lipopolysaccharide and flagellin. Multiple-factor analysis identify that the metabolic improvements induced by ω-3 PUFAs were accompanied by a reduced production of the proinflammatory cytokine TNFα, and that ω-3 PUFA supplementation had a stronger effect on improving the hepatic fatty acid profile than endogenous ω-3 PUFA. While endogenous ω-3 PUFA production preferably improves glucose tolerance and insulin resistance, ω-3 PUFA intake appears to be required to elicit selective changes in hepatic endocannabinoidome signaling that are essential to alleviate high-fat diet-induced hepatic steatosis.

Combining metabolomics and machine learning models as a tool to distinguish non-classic 21-hydroxylase deficiency from polycystic ovary syndrome without adrenocorticotropic hormone testing.

STUDY QUESTION: Can a combination of metabolomic signature and machine learning (ML) models distinguish nonclassic 21-hydroxylase deficiency (NC21OHD) from polycystic ovary syndrome (PCOS) without adrenocorticotrophic hormone (ACTH) testing? SUMMARY ANSWER: A single sampling methodology may be an alternative to the dynamic ACTH test in order to exclude the diagnosis of NC21OHD in the presence of a clinical hyperandrogenic presentation at any time of the menstrual cycle. WHAT IS KNOWN ALREADY: The clinical presentation of patients with NC21OHD is similar with that for other disorders of androgen excess. Currently, cosyntropin stimulation remains the gold standard diagnosis of NC21OHD. STUDY DESIGN, SIZE, DURATION: The study was designed using a bicentric recruitment: an internal training set included 19 women with NC21OHD and 19 controls used for developing the model; a test set included 17 NC21OHD, 72 controls and 266 PCOS patients used to evaluate the performance of the diagnostic strategy thanks to an ML approach. PARTICIPANTS/MATERIALS, SETTING, METHODS: Fifteen steroid species were measured in serum by liquid chromatography-mass spectrometry (LC-MS/MS). This set of 15 steroids (defined as 'steroidome') used to map the steroid biosynthesis pathway was the input for our models. MAIN RESULTS AND THE ROLE OF CHANCE: From a single sample, modeling involving metabolic pathway mapping by profiling 15 circulating steroids allowed us to identify perfectly NC21OHD from a confounding PCOS population. The constructed model using baseline LC-MS/MS-acquired steroid fingerprinting successfully excluded all 17 NC21OHDs (sensitivity and specificity of 100%) from 266 PCOS from an external testing cohort of originally 549 women, without the use of ACTH testing. Blood sampling timing during the menstrual cycle phase did not impact the efficiency of our model. LIMITATIONS, REASONS FOR CAUTION: The main limitations were the use of a restricted and fully prospective cohort as well as an analytical issue, as not all laboratories are equipped with mass spectrometers able to routinely measure this panel of 15 steroids. Moreover, the robustness of our model needs to be established with a larger prospective study for definitive validation in clinical practice. WIDER IMPLICATIONS OF THE FINDINGS: This tool makes it possible to propose a new semiology for the management of hyperandrogenism. The model presents better diagnostic performances compared to the current reference strategy. The management of patients may be facilitated by limiting the use of ACTH tests. Finally, the modeling process allows a classification of steroid contributions to rationalize the biomarker approach and highlight some underlying pathophysiological mechanisms. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by 'Agence Française de Lutte contre le dopage' and DIM Région Ile de France. This study was supported by the French institutional PHRC 2010-AOR10032 funding source and APHP. All authors declare no competing financial interests. TRIAL REGISTRATION NUMBER: N/A.

Adipocytes secretome from normal and tumor breast favor breast cancer invasion by metabolic reprogramming.

BACKGROUND: Adipose tissue is a major component of breast stroma. This study focused on delineating the effects of adipose stem cells (ASCs) derived from breast of healthy women and cancer patients with normal or tumor breast cells. METHODS: The ASCs were induced to differentiate into adipocytes, and the subsequent adipocyte conditioned media (ACM) were evaluated for their fatty acid profile, adipokine secretion and influence on proliferation, migration and invasion on tumoral (MCF-7 and SUM159) and normal (HMEC) human breast cell lines. RESULTS: An enrichment of arachidonic acid was observed in ACM from tumor tissues. Adipose tissues from tumor free secrete twice as much leptin than those from proximal or distal to the tumor. All ACMs display proliferative activity and favor invasiveness of SUM159 cells compared to MCF-7 and HMEC. All ACMs induced lipid droplets accumulation in MCF-7 cells and increased CD36 expression in tumor cells. CONCLUSION: We conclude that among secreted factors analyzed, only arachidonic acid and leptin levels did discriminate ASCs from tumor-bearing and tumor-free breasts emphasizing the importance that other cell types could contribute to the adipose tissue secretome in a tumor context.

High performance liquid chromatography-tandem mass spectrometry quantification of tryptophan metabolites in human serum and stool - Application to clinical cohorts in Inflammatory Bowel Diseases.

Tryptophan, an essential amino acid, and its metabolites are involved in many physiological processes including neuronal functions, immune system, and gut homeostasis. Alterations to tryptophan metabolism are associated with various pathologies such as neurologic, psychiatric disorders, inflammatory bowel diseases (IBD), metabolic disorders, and cancer. It is consequently critical to develop a reliable, quantitative method for the analysis of tryptophan and its downstream metabolites from the kynurenine, serotonin, and indoles pathways. An LC-MS/MS method was designed for the analysis of tryptophan and 20 of its metabolites, without derivatization and performed in a single run. This method was validated for both serum and stool. The comparisons between serum and plasma, collected with several differing anticoagulants, showed significant differences only for serotonin. References values were established in sera and stools from healthy donors. For stool samples, as a proof of concept, the developed method was applied to a healthy control group and an IBD patient group. Results showed significant differences in the concentrations of tryptophan, xanthurenic acid, kynurenic acid, indole-3-lactic acid, and picolinic acid. This method allowed an extensive analysis of the three tryptophan metabolic pathways in two compartments. Beyond the application to IBD patients, the clinical use of this method is wide-ranging and may be applied to other pathological conditions involving tryptophan metabolism, such as neurological, psychiatric, or auto-inflammatory pathologies.

Alteration of the gut microbiota's composition and metabolic output correlates with COVID-19-like severity in obese NASH hamsters.

Obese patientss with nonalcoholic steatohepatitis (NASH) are particularly prone to developing severe forms of coronavirus disease 19 (COVID-19). The gut-to-lung axis is critical during viral infections of the respiratory tract, and a change in the gut microbiota's composition might have a critical role in disease severity. Here, we investigated the consequences of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on the gut microbiota in the context of obesity and NASH. To this end, we set up a nutritional model of obesity with dyslipidemia and NASH in the golden hamster, a relevant preclinical model of COVID-19. Relative to lean non-NASH controls, obese NASH hamsters develop severe inflammation of the lungs and liver. 16S rRNA gene profiling showed that depending on the diet, SARS-CoV-2 infection induced various changes in the gut microbiota's composition. Changes were more prominent and transient at day 4 post-infection in lean animals, alterations still persisted at day 10 in obese NASH animals. A targeted, quantitative metabolomic analysis revealed changes in the gut microbiota's metabolic output, some of which were diet-specific and regulated over time. Our results showed that specifically diet-associated taxa are correlated with disease parameters. Correlations between infection variables and diet-associated taxa highlighted a number of potentially protective or harmful bacteria in SARS-CoV-2-infected hamsters. In particular, some taxa in obese NASH hamsters (e.g. Blautia and Peptococcus) were associated with pro-inflammatory parameters in both the lungs and the liver. These taxon profiles and their association with specific disease markers suggest that microbial patterns might influence COVID-19 outcomes.

Identification of bronchoalveolar and blood immune-inflammatory biomarker signature associated with poor 28-day outcome in critically ill COVID-19 patients.

The local immune-inflammatory response elicited by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is still poorly described, as well as the extent to which its characteristics may be associated with the outcome of critical Coronavirus disease 2019 (COVID-19). In this prospective monocenter study, all consecutive COVID-19 critically ill patients admitted from February to December 2020 and explored by fiberoptic bronchoscopy with bronchoalveolar lavage (BAL) were included. Biological assays, including digital ELISA cytokine profiling and targeted eicosanoid metabolomic analysis, were performed on paired blood and BAL fluid (BALF). Clinical outcome was assessed through the World Health Organization 10-point Clinical Progression Scale (WHO-CPS) at the 28th day (D28) following the admission to intensive care unit. A D28-WHO-CPS value higher than 5 defined a poor outcome. Seventy-six patients were included, 45 (59%) had a poor day-28 outcome. As compared to their counterparts, patients with D28-WHO-CPS > 5 exhibited a neutrophil-predominant bronchoalveolar phenotype, with a higher BALF neutrophil/lymphocyte ratio, a blunted local type I interferon response, a decompartimentalized immune-inflammatory response illustrated by lower BALF/blood ratio of concentrations of IL-6 (1.68 [0.30-4.41] vs. 9.53 [2.56-19.1]; p = 0.001), IL-10, IL-5, IL-22 and IFN-γ, and a biological profile of vascular endothelial injury illustrated by a higher blood concentration of VEGF and higher blood and/or BALF concentrations of several vasoactive eicosanoids. In critically ill COVID-19 patients, we identified bronchoalveolar and blood immune-inflammatory biomarker signature associated with poor 28-day outcome.

Fecal microbiota and bile acids in IBD patients undergoing screening for colorectal cancer.

Due to the potential role of the gut microbiota and bile acids in the pathogenesis of both inflammatory bowel disease (IBD) and sporadic colorectal cancer, we aimed to determine whether these factors were associated with colorectal cancer in IBD patients. 215 IBD patients and 51 non-IBD control subjects were enrolled from 10 French IBD centers between September 2011 and July 2018. Fecal samples were processed for bacterial 16S rRNA gene sequencing and bile acid profiling. Demographic, clinical, endoscopic, and histological outcomes were recorded. Characteristics of IBD patients included: median age: 41.6 (IQR 22); disease duration 13.2 (13.1); 47% female; 21.9% primary sclerosing cholangitis; 109 patients with Crohn's disease (CD); 106 patients with ulcerative colitis (UC). The prevalence of cancer was 2.8% (6/215: 1 CD; 5 UC), high-grade dysplasia 3.7% (8/215) and low-grade dysplasia 7.9% (17/215). Lachnospira was decreased in IBD patients with cancer, while Agathobacter was decreased and Escherichia-Shigella increased in UC patients with any neoplasia. Bile acids were not associated with cancer or neoplasia. Unsupervised clustering identified three gut microbiota clusters in IBD patients associated with bile acid composition and clinical features, including a higher risk of neoplasia in UC in two clusters when compared to the third (relative risk (RR) 4.07 (95% CI 1.6-10.3, P < .01) and 3.56 (95% CI 1.4-9.2, P < .01)). In this multicentre observational study, a limited number of taxa were associated with neoplasia and exploratory microbiota clusters co-associated with clinical features, including neoplasia risk in UC. Given the very small number of cancers, the robustness of these findings will require assessment and validation in future studies.

Analysis of a pitfall in congenital adrenal hyperplasia newborn screening: evidence of maternal use of corticoids detected on dried blood spot.

Neonatal screening for congenital adrenal hyperplasia (CAH) faces many specific challenges. It must be done using a performant analytical approach that combines sensitivity and specificity to capture the potential causes of mortality during the first week of life, such as salt wasting and glucocorticoid deficiency. Here, we confirm that maternal inhaled corticosteroid intake during pregnancy is a possible cause of missed CAH diagnosis. Thanks to liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis, we were able to quantify endogenous steroid metabolites and also detect the presence of exogenous steroids in the dried blood spot of a newborn. Adding LC-MS/MS analysis as second-tier test, especially one that includes both 17-hydroxyprogesterone and 21-deoxycortisol measurements, would probably improve CAH diagnosis. In familial neonatal screening one could also look for maternal corticosteroid therapies that are hidden to prevent false-negative tests.