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Adrenocortical carcinoma (ACC) is a very rare and aggressive malignant disease. Therefore, overall survival (OS) has long been considered as the best endpoint. Yet, a unique endpoint is not optimal to take into account the heterogeneity in tumor profile and the diversification of therapeutic option. The purpose of this mini review was to describe endpoints used in the past, present and future in the field of ACC.Pubmed and Clinicaltrial.gov were used to identify relevant studies.Before year 2000 only three endpoints were regularly used: OS, recurrence-free survival (RFS) and response rate. These endpoints were used because ACC was seen as a homogeneous diseases with a high recurrence rate and low rate of long-term survival. Since 2000; along with the apparition of new class of drug, progression-free survival (PFS) has been more and more used. Other endpoints as "time to chemotherapy" or "Progression-free survival 2" were used to evaluate multimodal therapies or treatment with a delayed action. Finally, there is a hope that in the near future, quality of life along with other patient-reported outcomes may be used more frequently.While OS and PFS are currently the most used endpoints in ACC, new endpoints are needed to better take into account the challenges offered by different situations and treatment strategies.

Therapies for metastatic SDHB-dependent pheochromocytoma and paraganglioma (PPGL) are limited and poorly efficient. New targeted therapies and identification of early non-invasive biomarkers of response are thus urgently needed for these patients. We characterized an in vivo allograft model of spontaneously immortalized murine chromaffin cells (imCC) with inactivation of the Sdhb gene by dynamic contrast-enhanced MRI (DCE-MRI) and 18FDG-PET. We evaluated the response to several therapies: IACS-010759 (mitochondrial respiratory chain complex I inhibitor), sunitinib (tyrosine kinase inhibitor with anti-angiogenic activity), talazoparib (poly ADP ribose polymerase (PARP) inhibitor) combined or not to temozolomide (alkylating agent), pharmacological inhibitors of HIF2a (PT2385 and PT2977 (belzutifan)) and molecular inactivation of HIF2a (imCC Sdhb−/− shHIF2a). Multimodal imaging was performed, including magnetic resonance spectroscopy (1H-MRS) to monitor the level of succinate in vivo. The allografted model of Sdhb−/− imCC reflected SDHB-deficient tumors, with increased angiogenesis and a particular avidity for 18FDG. After 14 days of treatment, IACS-010759, sunitinib and talazoparib at high doses allowed a significant reduction of the tumor volumes. In contrast to the tumor growth inhibition observed in Sdhb−/− shHIF2a imCC tumors, pharmacological inhibitors of HIF2a (PT2385 and belzutifan) showed no antitumor action in this model, alone or in combination with sunitinib. 1H-MRS, but not DCE-MRI, enabled the monitoring response to sunitinib, which was the best treatment in this study, promoting a decrease in succinate levels detected in vivo. This study paves the way for new therapeutic options and reveals a potential new early biomarker of response to treatment in SDHB-dependent PPGL.

International audience; La succinate déshydrogénase (SDH) est une enzyme mitochondriale qui participe au cycle de Krebs et à la chaîne respiratoire. Quand elles sont à l’origine de cancers, les mutations des gènes codant les différentes sous-unités de la SDH sont responsables d’une prédisposition aux phéochromocytomes et aux paragangliomes, et, plus rarement, aux tumeurs stromales gastro-intestinales ou au cancer du rein. Une diminution de l’activité de la SDH, non expliquée par la génétique, s’observe aussi dans certains cancers plus fréquents. Une des conséquences de l’inactivation de la SDH est la production excessive de son substrat, le succinate, qui joue un rôle d’oncométabolite en promouvant un statut pseudohypoxique et d’importants remaniements épigénétiques. La compréhension de l’oncogenèse liée à la succinate déshydrogénase permet aujourd’hui de développer des méthodes diagnostiques innovantes et d’envisager des thérapies ciblées pour la prise en charge des patients atteints.

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Succinate dehydrogenase is a key enzyme in the tricarboxylic acid cycle and the electron transport chain. All four subunits of succinate dehydrogenase are tumor suppressor genes predisposing to paraganglioma, but only mutations in the SDHB subunit are associated with increased risk of metastasis. Here we generated an Sdhd knockout chromaffin cell line and compared it with Sdhb-deficient cells. Both cell types exhibited similar SDH loss of function, metabolic adaptation, and succinate accumulation. In contrast, Sdhb−/− cells showed hallmarks of mesenchymal transition associated with increased DNA hypermethylation and a stronger pseudo-hypoxic phenotype compared with Sdhd−/− cells. Loss of SDHB specifically led to increased oxidative stress associated with dysregulated iron and copper homeostasis in the absence of NRF2 activation. High-dose ascorbate exacerbated the increase in mitochondrial reactive oxygen species, leading to cell death in Sdhb−/− cells. These data establish a mechanism linking oxidative stress to iron homeostasis that specifically occurs in Sdhb-deficient cells and may promote metastasis. They also highlight high-dose ascorbate as a promising therapeutic strategy for SDHB-related cancers.Significance:Loss of different succinate dehydrogenase subunits can lead to different cell and tumor phenotypes, linking stronger 2-OG–dependent dioxygenases inhibition, iron overload, and ROS accumulation following SDHB mutation.

Supplementary Table 2 Related to Figure 2. Differentially methylated gene-based features in Sdhb-/- versus Sdhd-/- imCCs

Pheochromocytomas and paragangliomas (PPGL) are rare neuroendocrine tumors arising from the adrenal medulla or extra-adrenal paraganglia. Around 40% of all cases are caused by a germline mutation in a susceptibility gene, half of which being found in an SDHx gene (SDHA, SDHB, SDHC, SDHD or SDHAF2). They encode the four subunits and assembly factor of succinate dehydrogenase (SDH), a mitochondrial enzyme involved both in the tricarboxylic acid cycle and electron transport chain. SDHx mutations lead to the accumulation of succinate, which acts as an oncometabolite by inhibiting iron(II) and alpha-ketoglutarate-dependent dioxygenases thereby regulating the cell’s hypoxic response and epigenetic processes. Moreover, SDHx mutations induce cell metabolic reprogramming and redox imbalance. Major discoveries in PPGL pathophysiology have been made since the initial discovery of SDHD gene mutations in 2000, improving the understanding of their biology and patient management. It indeed provides new opportunities for diagnostic tools and innovative therapeutic targets in order to improve the prognosis of patients affected by these rare tumors, in particular in the context of metastatic diseases associated with SDHB mutations. This review first describes an overview of the pathophysiology and then focuses on clinical implications of the epigenetic and metabolic reprogramming of SDH-deficient PPGL.

Succinate dehydrogenase (SDH) is a mitochondrial enzyme that participates in both the tricarboxylic acid cycle and the electron transport chain. Mutations in genes encoding SDH are responsible for a predisposition to pheochromocytomas and paragangliomas, and more rarely, to gastrointestinal stromal tumors or renal cell carcinomas. A decrease in SDH activity, not explained by genetics, has also been observed in more common cancers. One of the consequences of the inactivation of SDH is the excessive production of its substrate, succinate, which acts as an oncometabolite by promoting a pseudohypoxic status and an extensive epigenetic rearrangement. Understanding SDH-related oncogenesis now makes it possible to develop innovative diagnostic methods and to consider targeted therapies for the management of affected patients.Rôle de la succinate déshydrogénase dans le cancer.La succinate déshydrogénase (SDH) est une enzyme mitochondriale qui participe au cycle de Krebs et à la chaîne respiratoire. Quand elles sont à l’origine de cancers, les mutations des gènes codant les différentes sous-unités de la SDH sont responsables d’une prédisposition aux phéochromocytomes et aux paragangliomes, et, plus rarement, aux tumeurs stromales gastro-intestinales ou au cancer du rein. Une diminution de l’activité de la SDH, non expliquée par la génétique, s’observe aussi dans certains cancers plus fréquents. Une des conséquences de l’inactivation de la SDH est la production excessive de son substrat, le succinate, qui joue un rôle d’oncométabolite en promouvant un statut pseudohypoxique et d’importants remaniements épigénétiques. La compréhension de l’oncogenèse liée à la succinate déshydrogénase permet aujourd’hui de développer des méthodes diagnostiques innovantes et d’envisager des thérapies ciblées pour la prise en charge des patients atteints.

Neuroendocrine tumors constitute a heterogeneous group of tumors arising from hormone-secreting cells and are generally associated with a dysfunction of secretion. Pheochromocytoma (Pheo) is a neuroendocrine tumor that develops from chromaffin cells of the adrenal medulla, and is responsible for an excess of catecholamine secretion leading to severe clinical symptoms such as hypertension, elevated stroke risk and various cardiovascular complications. Surprisingly, while the hypersecretory activity of Pheo is well known to pathologists and clinicians, it has never been carefully explored at the cellular and molecular levels. In the present study, we have combined catecholamine secretion measurement by carbon fiber amperometry on human tumor cells directly cultured from freshly resected Pheos, with the analysis by mass spectrometry of the exocytotic proteins differentially expressed between the tumor and the matched adjacent non-tumor tissue. In most patients, catecholamine secretion recordings from single Pheo cells revealed a higher number of exocytic events per cell associated with faster kinetic parameters. Accordingly, we unravel significant tumor-associated modifications in the expression of key proteins involved in different steps of the calcium-regulated exocytic pathway. Altogether, our findings indicate that dysfunction of the calcium-regulated exocytosis at the level of individual Pheo cell is a cause of the tumor-associated hypersecretion of catecholamines.

Pheochromocytoma (Pheo) is a neuroendocrine tumor that develops from chromaffin cells of the adrenal medulla, and is responsible of an excess of catecholamines secretion leading to severe clinical symptoms such as hypertension, elevated stroke risk and various cardiovascular complications. Surprisingly, hypersecretory activity of Pheo has never been explored at the cellular and molecular levels from individual tumor cells. In the present study, we have combined catecholamine secretion measurement by carbon fiber amperometry on human tumor cells directly cultured from freshly resected Pheo, with the analysis by mass spectrometry of the exocytotic proteins differentially expressed between the tumor and the matched adjacent non-tumor tissue. Catecholamine secretion recordings from individual Pheo cells obtained from most patients reveal a higher number of exocytic events per cell associated with faster kinetic parameters. Accordingly, we unravel significant tumor-associated modifications in the expression of key proteins involved in different steps of the calcium-regulated exocytic pathway. Altogether, our findings indicate that dysfunction of the calcium-regulated exocytosis at the level of individual Pheo cell is a cause of the tumor-associated hypersecretion of catecholamines.

International audience; Increasingly common, neuroendocrine tumors (NETs) are regarded nowadays as neoplasms potentially causing debilitating symptoms and life-threatening medical conditions. Pheochromocytoma is a NET that develops from chromaffin cells of the adrenal medulla, and is responsible for an excessive secretion of catecholamines. Consequently, patients have an increased risk for clinical symptoms such as hypertension, elevated stroke risk and various cardiovascular complications. Somatostatin analogues are among the main anti-secretory medical drugs used in current clinical practice in patient with NETs. However, their impact on pheochromocytoma-associated catecholamine hypersecretion remains incompletely explored. This study investigated the potential efficacy of octreotide and pasireotide (SOM230) on human tumor cells directly cultured from freshly resected pheochromocytomas using an implemented catecholamine secretion measurement by carbon fiber amperometry. SOM230 treatment efficiently inhibited nicotine-induced catecholamine secretion both in bovine chromaffin cells and in human tumor cells whereas octreotide had no effect. Moreover, SOM230 specifically decreased the number of exocytic events by impairing the stimulation-evoked calcium influx as well as the nicotinic receptor-activated inward current in human pheochromocytoma cells. Altogether, our findings indicate that SOM230 acts as an inhibitor of catecholamine secretion through a mechanism involving the nicotinic receptor and might be considered as a potential anti-secretory treatment for patients with pheochromocytoma.

Succinate dehydrogenase is a key enzyme in the tricarboxylic acid cycle and the electron transport chain. All four subunits of succinate dehydrogenase are tumor suppressor genes predisposing to paraganglioma, but only mutations in the SDHB subunit are associated with increased risk of metastasis. Here we generated an Sdhd knockout chromaffin cell line and compared it with Sdhb-deficient cells. Both cell types exhibited similar SDH loss of function, metabolic adaptation, and succinate accumulation. In contrast, Sdhb−/− cells showed hallmarks of mesenchymal transition associated with increased DNA hypermethylation and a stronger pseudo-hypoxic phenotype compared with Sdhd−/− cells. Loss of SDHB specifically led to increased oxidative stress associated with dysregulated iron and copper homeostasis in the absence of NRF2 activation. High-dose ascorbate exacerbated the increase in mitochondrial reactive oxygen species, leading to cell death in Sdhb−/− cells. These data establish a mechanism linking oxidative stress to iron homeostasis that specifically occurs in Sdhb-deficient cells and may promote metastasis. They also highlight high-dose ascorbate as a promising therapeutic strategy for SDHB-related cancers. Significance: Loss of different succinate dehydrogenase subunits can lead to different cell and tumor phenotypes, linking stronger 2-OG–dependent dioxygenases inhibition, iron overload, and ROS accumulation following SDHB mutation.

Radioiodine treatment (RAI) represents the most widespread and effective therapy for differentiated thyroid cancer (DTC). RAI goals encompass ablative (destruction of thyroid remnants, to enhance thyroglobulin predictive value), adjuvant (destruction of microscopic disease to reduce recurrences), and therapeutic (in case of macroscopic iodine avid lesions) purposes, but its use has evolved over time. Randomized trial results have enabled the refinement of RAI indications, moving from a standardized practice to a tailored approach. In most cases, low-risk patients may safely avoid RAI, but where necessary, a simplified protocol, based on lower iodine activities and human recombinant TSH preparation, proved to be just as effective, reducing overtreatment or useless impairment of quality of life. In pediatric DTC, RAI treatments may allow tumor healing even at the advanced stages. Finally, new challenges have arisen with the advancement in redifferentiation protocols, through which RAI still represents a leading therapy, even in former iodine refractory cases. RAI therapy is usually well-tolerated at low activities rates, but some concerns exist concerning higher cumulative doses and long-term outcomes. Despite these achievements, several issues still need to be addressed in terms of RAI indications and protocols, heading toward the RAI strategy of the future.

Introduction Les progres dans la comprehension de l’oncogenese des pheochromocytomes et des paragangliomes metastatiques (PPGLm) mutes SDHB nous permettent d’envisager le recours a de nouvelles therapies ciblees dans cette pathologie rare dont le pronostic reste reserve. Materiel et methodes La reponse a plusieurs therapies a ete evaluee dans un modele in vivo d’allogreffes de cellules chromaffines murines spontanement immortalisees (imCC) avec inactivation du gene Sdhb. Differentes imageries multimodales ont ete realisees, dont une spectroscopie par resonnance magnetique permettant de monitorer le taux de succinate in vivo. Un modele d’allogreffes des cellules imCC Sdhb-/- avec inactivation de HIF2α (imCC Sdhb-/- shHIF2α) a egalement ete analyse. Resultats Apres 14 jours de traitement, l’ IACS (inhibiteur du complexe I de la chaine respiratoire mitochondriale), le Sunitinib (inhibiteur de tyrosine kinase a activite anti-angiogenique) et le Talazoparib (inhibiteur de PARP) a posologie elevee permettent une reduction significative du volume tumoral. Seuls les traitements par IACS et Sunitinib sont bien toleres. La combinaison du Talazoparib avec un agent alkylant, le Temozolomide, et la combinaison de l’IACS avec le Sunitinib potentialisent ces effets anti-tumoraux au prix de toxicites plus importantes. L’efficacite anti-tumorale du Sunitinib est correlee a une diminution du succinate in vivo. Contrairement a l’inhibition de la croissance tumorale observee dans les tumeurs imCC Sdhb-/- shHIF2α, les deux inhibiteurs pharmacologiques de HIF2α (PT2385 et PT2977) ne semblent avoir que peu d’action anti-tumorale dans ce modele. Conclusion Ces travaux preliminaires ouvrent la voie vers de nouvelles options therapeutiques dans les PPGLm SDHB-dependants.

Background & aims Cholangiopathies are chronic liver diseases in which damaged cholangiocytes trigger a proinflammatory and profibrotic reaction. The nuclear vitamin D receptor (VDR) is highly expressed in cholangiocytes and exerts immune-regulatory functions in these cells. In the present study, we examined the protective function of VDR and other vitamin D signaling pathways in chronic cholangiopathy and cholangiocytes. Methods Vdr was invalidated in Abcb4 knockout mice, a widely used animal model of chronic cholangiopathy. The impact of vitamin D signaling on cholangiopathy features was examined in vivo and in cholangiocytes (primary and cell lines). Results Cholangiopathy features (i.e, cholestasis, ductular reaction and fibrosis) were aggravated in Vdr;Abcb4 double knockout mice compared to the Abcb4 simple knockout, and associated with an overexpression of proinflammatory factors. The proinflammatory phenotype of cholangiocytes was also exacerbated following VDR silencing in vitro. The expression of proinflammatory factors and the severity of cholangiopathy were reduced in the double knockout mice treated with the vitamin D analog calcipotriol or with vitamin D. In vitro, the inflammatory response to TNFα was significantly reduced by calcipotriol in biliary cells silenced for VDR, and this effect was abolished by co-silencing the plasma membrane receptor of vitamin D, protein disulfide-isomerase A3 (PDIA3). Conclusions Our results demonstrate an anti-inflammatory role of VDR signaling in cholangiocytes and cholangiopathy. They also provide evidence for PDIA3-mediated anti-inflammatory effects of vitamin D and vitamin D analog in these settings.

Context The behavior of locally advanced pheochromocytoma (LAP) remains unknown. Objective We characterized the population with LAP and recurrence-free survival (RFS). Methods This retrospective multicentric study was run within the ENDOCAN-COMETE network and French Group of Endocrine Tumors (GTE) from 2003 to 2018, including patients from 11 French referral centers with LAP as defined by capsular invasion, vascular invasion, adipose tissue invasion, and/or positive locoregional lymph nodes at diagnosis without evidence of distant metastasis. The main outcome measure was recurrence, defined as tumor reappearance, including local site and/or distant metastasis. The primary endpoint was RFS analysis; secondary endpoints were characterization, overall survival (OS), and prognostic factors of recurrence. Results Among 950 patients, 90 (9%) exhibited LAP criteria and 55 met inclusion criteria (median age, 53 years; 61% males; 14% with germline mutation; 84% with catecholamine excess). LAP was defined by 31 (56%) capsular invasions, 27 (49%) fat invasions, 6 (11%) positive lymph nodes, and 22 (40%) vascular invasions. After median follow-up of 54 months (range, 6-180), 12 patients (22%) had recurrences and 3 (5%) died of metastatic disease. Median RFS was 115 months (range, 6-168). Recurrences were local in 2 patients, distant in 2, and both local and distant in 8 patients. Median OS of patients was not reached. Size above 6.5 cm (P = 0.019) and Ki-67 > 2% (P = 0.028) were identified as independent significant prognostic factors in multivariate analysis. Conclusion LAP represents 9% of pheochromocytoma’s population and has a metastatic behavior. This study paves the way for future pathological TNM classification.

BackgroundLaminopathies are a clinically heterogeneous group of disorders caused by mutations in the LMNA gene, which encodes the nuclear envelope proteins lamins A and C. The most frequent diseases associated with LMNA mutations are characterized by skeletal and cardiac involvement, and include autosomal dominant Emery-Dreifuss muscular dystrophy (EDMD), limb-girdle muscular dystrophy type 1B, and LMNA-related congenital muscular dystrophy (LMNA-CMD). Although the exact pathophysiological mechanisms responsible for LMNA-CMD are not yet understood, severe contracture and muscle atrophy suggest that impair skeletal muscle growth may contribute to the disease severity.MethodsWe used human muscle stem cells (MuSCs) carrying 4 different LMNA mutations and two mouse models of muscle laminopathies, representing a spectrum of disease severity, to investigate the ability of skeletal muscle to differentiate and to hypertrophy in response to mechanical challenges. We extended these finding to individuals with LMNA-related muscular dystrophy using muscle biopsies.ResultsIn vitro, we observe impaired myogenic differentiation with disorganized cadherin/β catenin adhesion complexes in MuSCs carrying LMNA-CMD. We show that skeletal muscle from Lmna-CMD mice is unable to hypertrophy in response to functional overload, due to defective accretion of activated MuSCs, defective protein synthesis and defective remodeling of the neuro-muscular junction. Moreover, stretched myotubes and overloaded muscle fibers with LMNA-CMD mutations display aberrant mechanical regulation of the Yes-Associated Protein (YAP), a key sensor and mediator of mechanical cues. We also observe defects in MuSC activation and YAP signaling in muscle biopsies from LMNA-CMD patients. These phenotypes are not recapitulated in closely-related EDMD models.ConclusionsCombining studies in vitro, in vivo and patient samples, we find that LMNA-CMD mutations interfere with mechano-signaling pathways in skeletal muscle, implicating defective skeletal muscle growth as a pathogenic contributor for the severity of LMNA-related muscular dystrophy.

Mutations in the LMNA gene, encoding the nuclear envelope A-type lamins, are responsible for muscular dystrophies, the most severe form being the LMNA-related congenital muscular dystrophy (L-CMD), with severe defects in myonucleus integrity. We previously reported that L-CMD mutations compromise the ability of muscle stem cells to modulate the yes-associated protein (YAP), a pivotal factor in mechanotransduction and myogenesis. Here, we investigated the intrinsic mechanisms by which lamins influence YAP subcellular distribution, by analyzing different conditions affecting the balance between nuclear import and export of YAP. In contrast to wild type (WT) cells, LMNADK32 mutations failed to exclude YAP from the nucleus and to inactivate its transcriptional activity at high cell density, despite activation of the Hippo pathway. Inhibiting nuclear pore import abolished YAP nuclear accumulation in confluent mutant cells, thus showing persistent nuclear import of YAP at cell confluence. YAP deregulation was also present in congenital myopathy related to nesprin-1 KASH mutation, but not in cells expressing the LMNAH222P mutation, the adult form of lamin-related muscle dystrophy with reduced nuclear deformability. In conclusion, our data showed that L-CMD mutations increased YAP nuclear localization via an increased nuclear import and implicated YAP as a pathogenic contributor in muscle dystrophies caused by nuclear envelop defects.

Idiopathic pulmonary fibrosis (IPF) is characterized by the deposition of excessive extracellular matrix and the destruction of lung parenchyma, resulting from an aberrant wound-healing response. Although IPF is often associated with an imbalance in protease activity, the mechanisms underlying the sustained repair mechanisms are not fully understood. Here, we addressed the role of the recently identified, membrane-anchored serine protease human airway trypsin-like protease (HAT). In the present study, we show that both HAT expression and activity were up-regulated in human IPF specimens. Next, adenoviral overexpression of HAT before bleomycin challenge attenuated lung injury as well as extracellular matrix deposition in the bleomycin-induced pulmonary fibrosis model. In vitro, HAT prevented specific fibrosis-associated responses in primary human pulmonary fibroblasts and induced the expression of mediators associated with the prostaglandin E2 pathway. Altogether, our findings suggested that HAT could have a protective role in IPF and other fibrotic lung disorders.-Menou, A., Flajolet, P., Duitmen, J., Justet, A., Moog, S., Jaillet, M., Tabèze, L., Solhonne, B., Garnier, M., Mal, H., Mordant, P., Castier, Y., Cazes, A., Sallenave, J.-M., Mailleux, A. A., Crestani, B. Human airway trypsin-like protease exerts potent, antifibrotic action in pulmonary fibrosis.

Emerin is a nuclear envelope protein that contributes to genome organization and cell mechanics. Through its N-terminal LAP2-emerin-MAN1 (LEM)-domain, emerin interacts with the DNA-binding protein barrier-to-autointegration (BAF). Emerin also binds to members of the linker of the nucleoskeleton and cytoskeleton (LINC) complex. Mutations in the gene encoding emerin are responsible for the majority of cases of X-linked Emery-Dreifuss muscular dystrophy (X-EDMD). Most of these mutations lead to an absence of emerin. A few missense and short deletion mutations in the disordered region of emerin are also associated with X-EDMD. More recently, missense and short deletion mutations P22L, ∆K37 and T43I were discovered in emerin LEM-domain, associated with isolated atrial cardiac defects (ACD). Here we reveal which defects, at both the molecular and cellular levels, are elicited by these LEM-domain mutations. Whereas K37 mutation impaired the correct folding of the LEM-domain, P22L and T43I had no impact on the 3D structure of emerin. Surprisingly, all three mutants bound to BAF, albeit with a weaker affinity in the case of K37. In human myofibroblasts derived from a patient&rsquo, s fibroblasts, emerin ∆K37 was correctly localized at the inner nuclear membrane, but was present at a significantly lower level, indicating that this mutant is abnormally degraded. Moreover, SUN2 was reduced, and these cells were defective in producing actin stress fibers when grown on a stiff substrate and after cyclic stretches. Altogether, our data suggest that the main effect of mutation K37 is to perturb emerin function within the LINC complex in response to mechanical stress.

Background Idiopathic pulmonary fibrosis is a devastating fibrotic diffuse parenchymal lung disorder that remains refractory to pharmacological therapies. Therefore, novel treatments are urgently required. CCAAT/enhancer binding protein delta (C/EBPδ) is a transcription factor that mediates critical cellular functions in pathophysiology and which was recently suggested to be a key regulatory component in IPF. The purpose of this study was to prove or refute the importance of C/EBPδ in pulmonary fibrosis. Methods Pulmonary fibrosis was induced by intranasal instillation of bleomycin into wild-type and C/EBPδ deficient mice. At different time intervals after bleomycin instillation, fibrosis was assessed by hydroxyproline analysis, histochemistry and q-PCR for fibrotic marker expression. Results C/EBPδ deficient mice developed pulmonary fibrosis to a similar degree as wildtype mice as evident from similar Ashcroft scores, hydroxyproline levels and expression levels of collagen, fibronectin and α-smooth muscle actin at both 14 and 21 days after bleomycin instillation. The resolution of fibrosis, assessed at 48 days after bleomycin instillation, was also similar in wildtype and C/EBPδ deficient mice. In line with the lack of effect of C/EBPδ on fibrosis progression/resolution, macrophage recruitment and/or differentiation were also not different in wildtype or C/EBPδ deficient mice. Conclusions Overall, C/EBPδ does not seem to affect bleomycin-induced experimental pulmonary fibrosis and we challenge the importance of C/EBPδ in pulmonary fibrosis. Relevance for patients This study shows that the transcription factor C/EBPδ does not play a major role in the development of pulmonary fibrosis. Pharmacological targeting of C/EBPδ is therefore not likely to have a beneficial effect for patients suffering from pulmonary fibrosis.

Loss-of-function mutations in the SDHB subunit of succinate dehydrogenase predispose patients to aggressive tumors characterized by pseudohypoxic and hypermethylator phenotypes. The mechanisms leading to DNA hypermethylation and its contribution to SDH-deficient cancers remain undemonstrated. We examine the genome-wide distribution of 5-methylcytosine and 5-hydroxymethylcytosine and their correlation with RNA expression in SDHB-deficient tumors and murine Sdhb

Glycerol is a clinical biomarker of lipolysis that is mainly produced by adipose tissues. Blood glycerol content increases in pathological conditions such as metabolic and cardiovascular diseases or cancer cachexia, but also in response to energetic stress such as physical exercise. Accurate glycerol monitoring is therefore important in a range of healthcare contexts. However, current methods available for the quantification of glycerol are expensive, time-consuming, and require the extraction of plasma from blood, from which blood glycerol content is then extrapolated. Here, we report the development of a new point-of-care glycerometer device, DietSee, based on a strip-type biosensor that enables the quantification of glycerol directly from whole blood in 6 s. The performance of the biosensor was first evaluated using buffer solutions and spiked human and mouse plasma samples, and its response was compared with that of the gold-standard colorimetric method. The results obtained using DietSee correlated strongly with those from the reference method and demonstrated a linear response to glycerol levels across a wide range of concentrations (40–750 μM) that were representative of those in the human body. Next, the biosensor was validated using spiked human blood samples over a range of 30–55% hematocrit; it also demonstrated a strong correlation with reference measurements under these conditions (R2 = 0.97). In addition, the biosensor was only minimally affected by a variety of potential interferents (endogenous and exogenous) and was highly stable in storage (more than 2 years when strips were stored dry at 4 °C). Finally, we investigated the application of the biosensor to real-time monitoring of lipolysis and found that the DietSee is well adapted for this purpose in both human and mouse samples. To conclude, the novel DietSee glycerometer is a sensitive, selective, and rapid tool that enables characterization of the metabolic status of an individual by measuring the glycerol concentration from a single fingertip blood drop.

Introduction La matrice extra-cellulaire (MEC) est une structure dynamique composee a plus de 90 % de collagenes, qui influence la progression tumorale et l’apparition de metastases. Son role dans l’agressivite des pheochromocytomes et paragangliomes (PPGL) est inconnu. Materiel et methodes Nous avons analyse le transcriptome de 188 PPGL collectes par le reseau COMETE et de cellules chromaffines murines immortalisees (imCC) porteuses d’une inactivation des genes Sdhb ou Sdhd. Nous avons egalement realise l’analyse proteomique de la MEC secretee par les cellules (matrisome) ainsi que des etudes fonctionnelles de migration et d’adhesion apres echanges de matrice entre les differents types cellulaires. Le role de la pseudo-hypoxie a ete etudie apres inactivation de Hif2a dans les imCC Sdhb-/-. Resultats Les genes codant les collagenes sont surexprimes dans les tumeurs mutees sur SDHB et les imCC Sdhb-/-. Cette surexpression est correlee aux niveaux proteiques observes dans le matrisome ou une augmentation majeure des collagenes est observee dans les imCC Sdhb-/- par rapport aux cellules natives (WT) et Sdhd-/-. La MEC produite par les imCC Sdhb-/- augmente l’adhesion et la migration des imCC WT et Sdhd-/-, suggerant son implication dans le phenotype metastatique associe a SDHB. L’inactivation de Hif2a permet une reversion partielle du phenotype des imCC Sdhb. Conclusion Ces resultats montrent que contrairement aux HIF prolyl-hydroxylases et au demethylases de l’ADN, les prolyl-hydroxylases necessaires a la maturation du collagene ne sont pas inhibees dans les PPGL SDHB. La reprogrammation de la MEC mediee par la pseudo-hypoxie pourrait participer au phenotype metastatique des PPGL SDHB.

Background: 18F-FDOPA positron emission tomography/computed tomography (PET/CT) is a sensitive nuclear imaging technology for the diagnosis of pheochromocytomas (PHEO). However, its utility in determining predictive factors for the secretion of catecholamines remains poorly studied. Methods: Thirty-nine histologically confirmed PHEO were included in this retrospective single-center study. Patients underwent 18F-FDOPA PET/CT before surgery, with an evaluation of several uptake parameters (standardized uptake values [SUVmax and SUVmean] and the metabolic burden [MB] calculated as follows: MB = SUVmean × tumor volume) and measurement of plasma and/or urinary metanephrine (MN), normetanephrine (NM), and chromogranin A. Thirty-five patients were screened for germline mutations in the RET, SDHx, and VHL genes. Once resected, primary cultures of 5 PHEO were used for real-time measurement of catecholamine release by carbon fiber amperometry. Results: The MB of the PHEO positively correlated with 24-h urinary excretion of NM (r = 0.64, p < 0.0001), MN (r = 0.49, p = 0.002), combined MN and NM (r = 0.75, p < 0.0001), and eventually plasma free levels of NM (r = 0.55, p = 0.006). In the mutated patients (3 SDHD, 2 SDHB, 3 NF1, 1 VHL, and 3 RET), a similar correlation was observed between MB and 24-h urinary combined MN and NM (r = 0.86, p = 0.0012). For the first time, we demonstrate a positive correlation between the PHEO-to-liver SUVmax ratio and the mean number of secretory granule fusion events of the corresponding PHEO cells revealed by amperometric spikes (p = 0.01). Conclusion: While the 18F-FDOPA PET/CT MB of PHEO strongly correlates with the concentration of MN, amperometric recordings suggest that 18F-FDOPA uptake could be enhanced by overactivity of catecholamine exocytosis.

Background Idiopathic pulmonary fibrosis is a devastating fibrotic diffuse parenchymal lung disorder that remains refractory to pharmacological therapies. Therefore, novel treatments are urgently required. CCAAT/enhancer binding protein delta (C/EBPδ) is a transcription factor that mediates critical cellular functions in pathophysiology and which was recently suggested to be a key regulatory component in IPF. The purpose of this study was to prove or refute the importance of C/EBPδ in pulmonary fibrosis. Methods Pulmonary fibrosis was induced by intranasal instillation of bleomycin into wild-type and C/EBPδ deficient mice. At different time intervals after bleomycin instillation, fibrosis was assessed by hydroxyproline analysis, histochemistry and q-PCR for fibrotic marker expression. Results C/EBPδ deficient mice developed pulmonary fibrosis to a similar degree as wildtype mice as evident from similar Ashcroft scores, hydroxyproline levels and expression levels of collagen, fibronectin and α-smooth muscle actin at both 14 and 21 days after bleomycin instillation. The resolution of fibrosis, assessed at 48 days after bleomycin instillation, was also similar in wildtype and C/EBPδ deficient mice. In line with the lack of effect of C/EBPδ on fibrosis progression/resolution, macrophage recruitment and/or differentiation were also not different in wildtype or C/EBPδ deficient mice. Conclusions Overall, C/EBPδ does not seem to affect bleomycin-induced experimental pulmonary fibrosis and we challenge the importance of C/EBPδ in pulmonary fibrosis. Relevance for patients This study shows that the transcription factor C/EBPδ does not play a major role in the development of pulmonary fibrosis. Pharmacological targeting of C/EBPδ is therefore not likely to have a beneficial effect for patients suffering from pulmonary fibrosis.

Adrenocortical lesions are characterized through imaging, hormonal and histopathological analysis. Our aim was to compare the radiological features of adrenocortical lesions with their cortisol-secreting status and histopathological Weiss score. Seventy five patients operated between 2004 and 2016 in the University Hospital of Nancy for either adrenocortical carcinomas (ACC) or adrenocortical adenomas (ACA) were enrolled in this study. We collected cortisol parameters, Computed Tomography (CT) scans (unenhanced density, wash-out (WO) analysis) and 18F-Fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) datas. The histopathological Weiss score ultimately differentiates ACA (score ≤ 2) from ACC (score ≥ 3). One-way ANOVA, Fisher’s exact and unpaired t tests were used for statistical analysis with significancy reached at p

Introduction: Human Airway Trypsin-like protease (HAT) belongs to the membrane-anchored serine proteases family and is expressed in the lung. While its pathophysiological functions remain unknown, HAT is involved in pulmonary remodeling processes such as asthma. In bronchial airways HAT stimulates fibroblast proliferation through Protease-Activated Receptor-2, which is instrumental in IPF. So far, the role of HAT in IPF has never been investigated. Methods: In controls or patients with IPF, HAT expression was analyzed in lung biopsies by immunohistochemistry, and by Western-blot (WB) in lung homogenates, Broncho-Alveolar Lavages (BAL) and primary pulmonary fibroblasts. In lung homogenates and BAL, HAT activity was measured, by the cleavage of a specific chromogenic substrate. In vitro, the effects of HAT on fibrotic markers expression, migration and proliferation were assessed in human primary pulmonary fibroblasts using WB, WST-1 and Boyden chamber assays. Results: HAT is increased in IPF lungs in epithelial cells, (myo)fibroblasts and macrophages. Accordingly, HAT expression and activity are enhanced in lung and BAL from IPF patients. In fibroblasts, HAT induces phosphorylation of kinases such as ERK, decreases fibronectine and collagen expression, strongly impairs migration. Moreover, HAT does not increase pulmonary fibroblasts proliferation. Conclusions: Unlike its deleterious role in chronic inflammatory disease, in IPF, HAT induces anti-fibrotic responses in vitro. Its deregulated expression in patients with IPF may constitute an insufficient mechanism of protection. Although further studies are warranted in HAT-deficient mice, our results suggest that an induction of HAT in IPF could be beneficial.

Introduction: Human Airway Trypsin-like protease (HAT) belongs to the membrane-anchored serine proteases family. While its physiological functions remain unknown, HAT is involved in pulmonary remodeling processes like asthma. In airways HAT stimulates fibroblast proliferation thought Protease-Activated Receptor-2, a pro-fibrotic receptor in IPF, but its role in IPF has never been investigated so far. Methods: In controls or patients with IPF, HAT expression was analysed in lung biopsies by immunohistochemistry and by Western-blot (WB) in lung homogenates, Broncho-Alveolar Lavages (BAL) and primary pulmonary fibroblasts. In homogenates and BAL, HAT activity was measured with a specific chromogenic substrate. In vitro , the effects of HAT on fibrotic markers expression, migration and proliferation were assessed in human primary pulmonary fibroblasts using WB, WST-1 and Boyden chamber assays. HAT expression and activity were measured in a mouse model of bleomycin-induced lung fibrosis. Results: HAT is prominent in IPF lungs in epithelial cells and (myo)fibroblasts. HAT expression and activity are enhanced in the lung and BAL from IPF patients. Results were similar in fibrotic mice. In fibroblast s, HAT induces decreases fibronectine and collagen expression, does not increase pulmonary fibroblasts proliferation and strongly decreases migration. Conclusions: In IPF, contrary to its deleterious role in chronic inflammatory disease, HAT induces anti-fibrotic responses in vitro . Its deregulated expression in IPF could constitute an insufficient mechanism of protection. These results should be confirmed in HAT-deficient mice. Our results suggest that an induction of HAT in IPF could be beneficial.