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1.
Lab Invest ; 104(9): 102122, 2024 Aug 02.
Article de Anglais | MEDLINE | ID: mdl-39098628

RÉSUMÉ

The assessment of chemotherapy response in osteosarcoma (OS), based on the average percentage of viable cells, is limited, as it overlooks the spatial heterogeneity of tumor cell response (foci of resistant cells), immune microenvironment, and bone microarchitecture. Despite the resulting positive classification for response to chemotherapy, some patients experience early metastatic recurrence, demonstrating that our conventional tools for evaluating treatment response are insufficient. We studied the interactions between tumor cells, immune cells (lymphocytes, histiocytes, and osteoclasts), and bone extracellular matrix (ECM) in 18 surgical resection samples of OS using multiplex and conventional immunohistochemistry (IHC: CD8, CD163, CD68, and SATB2), combined with multiscale characterization approaches in territories of good and poor response (GRT/PRT) to treatment. GRT and PRT were defined as subregions with <10% and ≥10% of viable tumor cells, respectively. Local correlations between bone ECM porosity and density of immune cells were assessed in these territories. Immune cell density was then correlated to overall patient survival. Two patterns were identified for histiocytes and osteoclasts. In poor responder patients, CD68 osteoclast density exceeded that of CD163 histiocytes but was not related to bone ECM load. Conversely, in good responder patients, CD163 histiocytes were more numerous than CD68 osteoclasts. For both of them, a significant negative local correlation with bone ECM porosity was found (P < ,01). Moreover, in PRT, multinucleated osteoclasts were rounded and intermingled with tumor cells, whereas in GRT, they were elongated and found in close contact with bone trabeculae. CD8 levels were always low in metastatic patients, and those initially considered good responders rapidly died from their disease. The specific recruitment of histiocytes and osteoclasts within the bone ECM, and the level of CD8 represent new features of OS response to treatment. The associated prognostic signatures should be integrated into the therapeutic stratification algorithm of patients after surgery.

2.
J Ovarian Res ; 17(1): 134, 2024 Jun 28.
Article de Anglais | MEDLINE | ID: mdl-38943138

RÉSUMÉ

BACKGROUND: Industrial progress has led to the omnipresence of chemicals in the environment of the general population, including reproductive-aged and pregnant women. The reproductive function of females is a well-known target of endocrine-disrupting chemicals. This function holds biological processes that are decisive for the fertility of women themselves and for the health of future generations. However, insufficient research has evaluated the risk of combined mixtures on this function. This study aimed to assess the direct impacts of a realistic exposure to eight combined environmental toxicants on the critical process of folliculogenesis. METHODS: Female rabbits were exposed daily and orally to either a mixture of eight environmental toxicants (F group) or the solvent mixture (NE group, control) from 2 to 19 weeks of age. The doses were computed from previous toxicokinetic data to reproduce steady-state serum concentrations in rabbits in the range of those encountered in pregnant women. Ovarian function was evaluated through macroscopic and histological analysis of the ovaries, serum hormonal assays and analysis of the expression of steroidogenic enzymes. Cellular dynamics in the ovary were further investigated with Ki67 staining and TUNEL assays. RESULTS: F rabbits grew similarly as NE rabbits but exhibited higher total and high-density lipoprotein (HDL) cholesterol levels in adulthood. They also presented a significantly elevated serum testosterone concentrations, while estradiol, progesterone, AMH and DHEA levels remained unaffected. The measurement of gonadotropins, androstenedione, pregnenolone and estrone levels yielded values below the limit of quantification. Among the 7 steroidogenic enzymes tested, an isolated higher expression of Cyp19a1 was measured in F rabbits ovaries. Those ovaries presented a significantly greater density/number of antral and atretic follicles and larger antral follicles without any changes in cellular proliferation or DNA fragmentation. No difference was found regarding the count of other follicle stages notably the primordial stage, the corpora lutea or AMH serum levels. CONCLUSION: Folliculogenesis and steroidogenesis seem to be subtly altered by exposure to a human-like mixture of environmental toxicants. The antral follicle growth appears promoted by the mixture of chemicals both in their number and size, potentially explaining the increase in atretic antral follicles. Reassuringly, the ovarian reserve estimated through primordial follicles number/density and AMH is spared from any alteration. The consequences of these changes on fertility and progeny health have yet to be investigated.


Sujet(s)
Follicule ovarique , Réserve ovarienne , Femelle , Animaux , Lapins , Follicule ovarique/effets des médicaments et des substances chimiques , Follicule ovarique/métabolisme , Humains , Réserve ovarienne/effets des médicaments et des substances chimiques , Perturbateurs endocriniens/toxicité , Polluants environnementaux/toxicité , Ovaire/effets des médicaments et des substances chimiques , Ovaire/métabolisme , Exposition environnementale/effets indésirables
3.
Nat Commun ; 14(1): 7112, 2023 11 06.
Article de Anglais | MEDLINE | ID: mdl-37932311

RÉSUMÉ

An unresolved issue in contemporary biomedicine is the overwhelming number and diversity of complex images that require annotation, analysis and interpretation. Recent advances in Deep Learning have revolutionized the field of computer vision, creating algorithms that compete with human experts in image segmentation tasks. However, these frameworks require large human-annotated datasets for training and the resulting "black box" models are difficult to interpret. In this study, we introduce Kartezio, a modular Cartesian Genetic Programming-based computational strategy that generates fully transparent and easily interpretable image processing pipelines by iteratively assembling and parameterizing computer vision functions. The pipelines thus generated exhibit comparable precision to state-of-the-art Deep Learning approaches on instance segmentation tasks, while requiring drastically smaller training datasets. This Few-Shot Learning method confers tremendous flexibility, speed, and functionality to this approach. We then deploy Kartezio to solve a series of semantic and instance segmentation problems, and demonstrate its utility across diverse images ranging from multiplexed tissue histopathology images to high resolution microscopy images. While the flexibility, robustness and practical utility of Kartezio make this fully explicable evolutionary designer a potential game-changer in the field of biomedical image processing, Kartezio remains complementary and potentially auxiliary to mainstream Deep Learning approaches.


Sujet(s)
Algorithmes , Traitement d'image par ordinateur , Humains , Traitement d'image par ordinateur/méthodes , Microscopie , Évolution biologique , Sémantique
4.
J Immunother Cancer ; 11(10)2023 10.
Article de Anglais | MEDLINE | ID: mdl-37899130

RÉSUMÉ

BACKGROUND: Follicular lymphoma (FL), the most common indolent non-Hodgkin's Lymphoma, is a heterogeneous disease and a paradigm of the contribution of immune tumor microenvironment to disease onset, progression, and therapy resistance. Patient-derived models are scarce and fail to reproduce immune phenotypes and therapeutic responses. METHODS: To capture disease heterogeneity and microenvironment cues, we developed a patient-derived lymphoma spheroid (FL-PDLS) model culturing FL cells from lymph nodes (LN) with an optimized cytokine cocktail that mimics LN stimuli and maintains tumor cell viability. RESULTS: FL-PDLS, mainly composed of tumor B cells (60% on average) and autologous T cells (13% CD4 and 3% CD8 on average, respectively), rapidly organizes into patient-specific three-dimensional (3D) structures of three different morphotypes according to 3D imaging analysis. RNAseq analysis indicates that FL-PDLS reproduces FL hallmarks with the overexpression of cell cycle, BCR, or mTOR signaling related gene sets. FL-PDLS also recapitulates the exhausted immune phenotype typical of FL-LN, including expression of BTLA, TIGIT, PD-1, TIM-3, CD39 and CD73 on CD3+ T cells. These features render FL-PDLS an amenable system for immunotherapy testing. With this aim, we demonstrate that the combination of obinutuzumab (anti-CD20) and nivolumab (anti-PD1) reduces tumor load in a significant proportion of FL-PDLS. Interestingly, B cell depletion inversely correlates with the percentage of CD8+ cells positive for PD-1 and TIM-3. CONCLUSIONS: In summary, FL-PDLS is a robust patient-derived 3D system that can be used as a tool to mimic FL pathology and to test novel immunotherapeutic approaches in a context of personalized medicine.


Sujet(s)
Lymphome folliculaire , Humains , Lymphome folliculaire/traitement médicamenteux , Lymphome folliculaire/génétique , Récepteur cellulaire-2 du virus de l'hépatite A , Récepteur-1 de mort cellulaire programmée/métabolisme , Microenvironnement tumoral , Médecine de précision
5.
Toxics ; 11(5)2023 May 03.
Article de Anglais | MEDLINE | ID: mdl-37235240

RÉSUMÉ

Animal toxicological studies often fail to mimic the complexity of the human exposome, associating low doses, combined molecules and long-term exposure. Since the reproductive potential of a woman begins in the fetal ovary, the literature regarding the disruption of its reproductive health by environmental toxicants remains limited. Studies draw attention to follicle development, a major determinant for the quality of the oocyte, and the preimplantation embryo, as both of them are targets for epigenetic reprogramming. The "Folliculogenesis and Embryo Development EXPOsure to a mixture of toxicants: evaluation in the rabbit model" (FEDEXPO) project emerged from consideration of these limitations and aims to evaluate in the rabbit model the impacts of an exposure to a mixture of known and suspected endocrine disrupting chemicals (EDCs) during two specific windows, including folliculogenesis and preimplantation embryo development. The mixture combines eight environmental toxicants, namely perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), dichlorodiphenyldichloroethylene (DDE), hexachlorobenzene (HCB), ß-hexachlorocyclohexane (ß-HCH), 2,2'4,4'-tetrabromodiphenyl ether (BDE-47), di(2-ethylhexyl) phthalate (DEHP) and bisphenol S (BPS), at relevant exposure levels for reproductive-aged women based on biomonitoring data. The project will be organized in order to assess the consequences of this exposure on the ovarian function of the directly exposed F0 females and monitor the development and health of the F1 offspring from the preimplantation stage. Emphasis will be made on the reproductive health of the offspring. Lastly, this multigenerational study will also tackle potential mechanisms for the inheritance of health disruption via the oocyte or the preimplantation embryo.

6.
Cancers (Basel) ; 15(4)2023 Feb 04.
Article de Anglais | MEDLINE | ID: mdl-36831348

RÉSUMÉ

Background-The purpose of this study was to investigate the bone resorption, as well as the vascular and immune microenvironment, of jaw osteosarcomas (JO) and to correlate these features with patient clinical outcomes. Methods-We studied 50 JO biopsy samples by immunohistochemical analysis of tissue microarrays (TMAs). We investigated the bone remodeling markers RANK/RANKL/OPG, the endothelial glycoprotein CD146, and biomarkers of the immune environment (CD163 and CD68 of macrophages, CD4+ and CD8+ of tumor-infiltrating lymphocytes (TILs), and an immune checkpoint PD-1/PD-L1). The biomarkers were analyzed for their influence on progression (recurrence and metastasis), overall survival (OS), and disease-free survival (DFS). Results-A strong and significant correlation has been found between CD163 staining and lower OS and DFS. The level of CD4+ and CD8+ staining was low and non-significantly associated with survival outcomes. High levels of RANK and RANKL were found in the tumor samples and correlated with lower DFS. Conclusion-Our findings suggest that CD163+ TAMs represent markers of poor prognosis in JO. Targeting TAMs could represent a valuable therapeutic strategy in JO.

7.
Sci Adv ; 8(7): eabk3234, 2022 02 18.
Article de Anglais | MEDLINE | ID: mdl-35171665

RÉSUMÉ

Human cytotoxic T lymphocytes (CTLs) exhibit ultrarapid lytic granule secretion, but whether melanoma cells mobilize defense mechanisms with commensurate rapidity remains unknown. We used single-cell time-lapse microscopy to offer high spatiotemporal resolution analyses of subcellular events in melanoma cells upon CTL attack. Target cell perforation initiated an intracellular Ca2+ wave that propagated outward from the synapse within milliseconds and triggered lysosomal mobilization to the synapse, facilitating membrane repair and conferring resistance to CTL induced cytotoxicity. Inhibition of Ca2+ flux and silencing of synaptotagmin VII limited synaptic lysosomal exposure and enhanced cytotoxicity. Multiplexed immunohistochemistry of patient melanoma nodules combined with automated image analysis showed that melanoma cells facing CD8+ CTLs in the tumor periphery or peritumoral area exhibited significant lysosomal enrichment. Our results identified synaptic Ca2+ entry as the definitive trigger for lysosomal deployment to the synapse upon CTL attack and highlighted an unpredicted defensive topology of lysosome distribution in melanoma nodules.


Sujet(s)
Antinéoplasiques , Mélanome , Lymphocytes T CD8+ , Cytotoxicité immunologique , Humains , Lysosomes/métabolisme , Mélanome/métabolisme , Lymphocytes T cytotoxiques
9.
Hum Pathol ; 121: 46-55, 2022 03.
Article de Anglais | MEDLINE | ID: mdl-34995674

RÉSUMÉ

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can cause severe placental lesions leading rapidly to intrauterine fetal death (IUFD). From August 2020 to September 2021, in the pathology department of Toulouse Oncopole, we analyzed 50 placentas from COVID-19-positive unvaccinated mothers. The purpose of our study is to describe the clinicopathological characteristics of these placental damages and to understand the pathophysiology. Ten of them (20%) showed placental lesions with positive immunohistochemistry for SARS-CoV-2 in villous trophoblasts. In five cases (10%), we observed massive placental damage associating trophoblastic necrosis, fibrinous deposits, intervillositis, as well as extensive hemorrhagic changes due to SARS-CoV-2 infection probably responsible of IUFD by functional placental insufficiency. In five other cases, we found similar placental lesions but with a focal distribution that did not lead to IUFD but live birth. These lesions are independent of maternal clinical severity of COVID-19 infection because they occur despite mild maternal symptoms and are therefore difficult to predict. In our cases, they occurred 1-3 weeks after positive SARS-CoV-2 maternal real-time polymerase chain reaction testing and were observed in the 2nd and 3rd trimesters of pregnancies. When these lesions are focal, they do not lead to IUFD and can be involved in intrauterine growth restriction. Our findings, together with recent observations, suggest that future pregnancy guidance should include stricter pandemic precautions such as screening for a wider array of COVID-19 symptoms, enhanced ultrasound monitoring, as well as newborn medical surveillance.


Sujet(s)
COVID-19 , Complications infectieuses de la grossesse , COVID-19/complications , Femelle , Mort foetale/étiologie , Humains , Nouveau-né , Placenta/anatomopathologie , Grossesse , Complications infectieuses de la grossesse/anatomopathologie , SARS-CoV-2
10.
Cancer Lett ; 526: 112-130, 2022 02 01.
Article de Anglais | MEDLINE | ID: mdl-34826547

RÉSUMÉ

The cytoskeleton and cell-matrix adhesions constitute a dynamic network that controls cellular behavior during development and cancer. The Focal Adhesion Kinase (FAK) is a central actor of these cell dynamics, promoting cell-matrix adhesion turnover and active membrane fluctuations. However, the initial steps leading to FAK activation and subsequent promotion of cell dynamics remain elusive. Here, we report that the serine/threonine kinase PKCθ participates in the initial steps of FAK activation. PKCθ, which is strongly expressed in aggressive human breast cancers, controls the dynamics of cell-matrix adhesions and active protrusions through direct FAK activation, thereby promoting cell invasion and lung metastases. Using various tools for in vitro and live cell studies, we precisely decipher the molecular mechanisms of FAK activation. PKCθ directly interacts with the FAK FERM domain to open FAK conformation through PKCθ's specific V3 domain, while phosphorylating FAK at newly identified serine/threonine residues within nascent adhesions, inducing cell dynamics and aggressive behavior. This study thus places PKCθ-directed FAK opening and phosphorylations as an original mechanism controlling dynamic, migratory, and invasive abilities of aggressive breast cancer cells, further strengthening the emerging oncogenic function of PKCθ.


Sujet(s)
Tumeurs du sein/physiopathologie , Cytosquelette/métabolisme , Focal adhesion kinase 1/métabolisme , Protein Kinase C-theta/métabolisme , Protein-Serine-Threonine Kinases/métabolisme , Pseudopodes/métabolisme , Animaux , Adhérence cellulaire/physiologie , Lignée cellulaire tumorale , Mouvement cellulaire/physiologie , Femelle , Hétérogreffes , Humains , Cellules MCF-7 , Souris , Souris de lignée BALB C , Souris nude , Phosphorylation
11.
Amyloid ; 28(3): 153-157, 2021 Sep.
Article de Anglais | MEDLINE | ID: mdl-33583309

RÉSUMÉ

BACKGROUND: Congo red-positive material was described in normal and diseased parathyroids (adenoma and hyperplasia) 50 years ago. However, the incidence and the clinical significance of such observation are unknown, and the causal fibril protein has never been convincingly demonstrated. METHODS: We conducted the present study including an exceptional case report accompanied with a retrospective study of 105 parathyroid adenomas. We used histopathological, immunohistochemical, ultrastructural, mass spectrometry-based proteomic analysis of parathyroid adenoma tissue samples, and genetic analysis. RESULTS: We describe a 57-year-old man with mild hypercalcemia and elevated parathyroid hormone (PTH) level for whom histopathological analysis revealed a parathyroid adenoma associated with nodular typical amyloid deposits. Tandem mass spectrometry after laser microdissection (LMD-MS) of amyloid adenoma identified PTH as the fibril protein, and no germline mutation in the PTH gene was detected. Congo red-positive PTH-deposits were further observed in 6.6% of the parathyroid adenomas analyzed, and were associated with complete/incomplete or absent universal amyloid signature, but with fibrillar morphology at ultrastructural level. CONCLUSIONS: Inappropriate PTH production leads to progressive disease-amyloid aggregation of PTH in a subset of parathyroid adenomas, providing new insights into the pathophysiology of this condition and adding PTH to the list of amyloid protein derived from hormones.


Sujet(s)
Tumeurs de la parathyroïde , Amyloïde , Humains , Mâle , Adulte d'âge moyen , Hormone parathyroïdienne , Protéomique , Études rétrospectives , Spectrométrie de masse en tandem
12.
Oncoimmunology ; 7(10): e1486950, 2018.
Article de Anglais | MEDLINE | ID: mdl-30288350

RÉSUMÉ

Despite recent therapeutic progress, plasmablastic lymphoma (PBL), a distinct entity of high grade B cell lymphoma, is still an aggressive lymphoma with adverse prognosis. PBL commonly occurs in patients with HIV infection and PBL cells frequently express Epstein Barr virus (EBV) genome with type I latency. Occasionally however, PBL may develop in patients with an immunodepressed status without EBV and HIV infection. The aim of this study was to determine which PBL patients may benefit from the emerging strategies of immune checkpoint blockade. Here, we produced and analyzed the transcriptomic profiles of such tumors to address this question. Unsupervised hierarchical clustering analysis of PBL samples revealed they segregate according to their tumor EBV-status. Moreover, EBV+ PBL displays abundant leucocyte infiltrates and T-cell activation signatures, together with high expression levels of mRNA and protein markers of immune escape. This suggests that EBV infection induce an anti-viral cytotoxic immunity which progressively exhausts T lymphocytes and promotes the tolerogenic microenvironment of PBL. Hence, most EBV+ PBL patients presenting an early stage of cancer immune-editing process appear as the most eligible patients for immune checkpoint blockade therapies.

13.
Exp Neurol ; 291: 106-119, 2017 05.
Article de Anglais | MEDLINE | ID: mdl-28189729

RÉSUMÉ

Slc17a5-/- mice represent an animal model for the infantile form of sialic acid storage disease (SASD). We analyzed genetic and histological time-course expression of myelin and oligodendrocyte (OL) lineage markers in different parts of the CNS, and related this to postnatal neurobehavioral development in these mice. Sialin-deficient mice display a distinct spatiotemporal pattern of sialic acid storage, CNS hypomyelination and leukoencephalopathy. Whereas few genes are differentially expressed in the perinatal stage (p0), microarray analysis revealed increased differential gene expression in later postnatal stages (p10-p18). This included progressive upregulation of neuroinflammatory genes, as well as continuous down-regulation of genes that encode myelin constituents and typical OL lineage markers. Age-related histopathological analysis indicates that initial myelination occurs normally in hindbrain regions, but progression to more frontal areas is affected in Slc17a5-/- mice. This course of progressive leukoencephalopathy and CNS hypomyelination delays neurobehavioral development in sialin-deficient mice. Slc17a5-/- mice successfully achieve early neurobehavioral milestones, but exhibit progressive delay of later-stage sensory and motor milestones. The present findings may contribute to further understanding of the processes of CNS myelination as well as help to develop therapeutic strategies for SASD and other myelination disorders.


Sujet(s)
Encéphale/anatomopathologie , Régulation de l'expression des gènes au cours du développement/génétique , Leucoencéphalopathies , Troubles mentaux/étiologie , Transporteurs d'anions organiques/déficit , Maladie de surcharge en acide sialique , Symporteurs/déficit , Facteurs âges , Animaux , Animaux nouveau-nés , Encéphale/métabolisme , Incapacités de développement/étiologie , Incapacités de développement/génétique , Modèles animaux de maladie humaine , Protéine gliofibrillaire acide/métabolisme , Filaments intermédiaires/métabolisme , Leucoencéphalopathies/complications , Leucoencéphalopathies/étiologie , Leucoencéphalopathies/génétique , Protéine de membrane-1 associée au lysosome/métabolisme , Souris , Souris de lignée C57BL , Souris transgéniques , Transporteurs d'anions organiques/génétique , Maladie de surcharge en acide sialique/complications , Maladie de surcharge en acide sialique/génétique , Maladie de surcharge en acide sialique/anatomopathologie , Symporteurs/génétique
14.
PLoS One ; 11(9): e0161441, 2016.
Article de Anglais | MEDLINE | ID: mdl-27598321

RÉSUMÉ

The in vivo cutaneous nerve regeneration model using capsaicin is applied extensively to study the regenerative mechanisms and therapeutic efficacy of disease modifying molecules for small fiber neuropathy (SFN). Since mismatches between functional and morphological nerve fiber recovery are described for this model, we aimed at determining the capability of the capsaicin model to truly mimic the morphological manifestations of SFN in diabetes. As nerve and blood vessel growth and regenerative capacities are defective in diabetes, we focused on studying the key regulator of these processes, the neuropilin-1 (NRP-1)/semaphorin pathway. This led us to the evaluation of NRP-1 receptor expression in epidermis and dermis of subjects presenting experimentally induced small fiber neuropathy, diabetic polyneuropathy and of diabetic subjects without clinical signs of small fiber neuropathy. The NRP-1 receptor was co-stained with CD31 vessel-marker using immunofluorescence and analyzed with Definiens® technology. This study indicates that capsaicin application results in significant loss of epidermal NRP-1 receptor expression, whereas diabetic subjects presenting small fiber neuropathy show full epidermal NRP-1 expression in contrast to the basal expression pattern seen in healthy controls. Capsaicin induced a decrease in dermal non-vascular NRP-1 receptor expression which did not appear in diabetic polyneuropathy. We can conclude that the capsaicin model does not mimic diabetic neuropathy related changes for cutaneous NRP-1 receptor expression. In addition, our data suggest that NRP-1 might play an important role in epidermal nerve fiber loss and/or defective regeneration and that NRP-1 receptor could change the epidermal environment to a nerve fiber repellant bed possibly through Sem3A in diabetes.


Sujet(s)
Complications du diabète/génétique , Neuropathies diabétiques/génétique , Neuropiline 1/biosynthèse , Peau/métabolisme , Neuropathie des petites fibres/génétique , Adulte , Sujet âgé , Biopsie , Capsaïcine/métabolisme , Complications du diabète/anatomopathologie , Neuropathies diabétiques/anatomopathologie , Épiderme/métabolisme , Épiderme/anatomopathologie , Régulation de l'expression des gènes , Humains , Adulte d'âge moyen , Neurofibres/métabolisme , Neurofibres/anatomopathologie , Régénération nerveuse/génétique , Neuropiline 1/génétique , Antigènes CD31/génétique , Peau/anatomopathologie , Neuropathie des petites fibres/anatomopathologie
15.
BMC Res Notes ; 9: 280, 2016 May 23.
Article de Anglais | MEDLINE | ID: mdl-27215701

RÉSUMÉ

BACKGROUND: In this study we explored the possibility of automating the PGP9.5 immunofluorescence staining assay for the diagnosis of small fiber neuropathy using skin punch biopsies. The laboratory developed test (LDT) was subjected to a validation strategy as required by good laboratory practice guidelines and compared to the well-established gold standard method approved by the European Federation of Neurological Societies (EFNS). To facilitate automation, the use of thinner sections. (16 µm) was evaluated. Biopsies from previously published studies were used. The aim was to evaluate the diagnostic performance of the LDT compared to the gold standard. We focused on technical aspects to reach high-quality standardization of the PGP9.5 assay and finally evaluate its potential for use in large scale batch testing. RESULTS: We first studied linear nerve fiber densities in skin of healthy volunteers to establish reference ranges, and compared our LDT using the modifications to the EFNS counting rule to the gold standard in visualizing and quantifying the epidermal nerve fiber network. As the LDT requires the use of 16 µm tissue sections, a higher incidence of intra-epidermal nerve fiber fragments and a lower incidence of secondary branches were detected. Nevertheless, the LDT showed excellent concordance with the gold standard method. Next, the diagnostic performance and yield of the LDT were explored and challenged to the gold standard using skin punch biopsies of capsaicin treated subjects, and patients with diabetic polyneuropathy. The LDT reached good agreement with the gold standard in identifying small fiber neuropathy. The reduction of section thickness from 50 to 16 µm resulted in a significantly lower visualization of the three-dimensional epidermal nerve fiber network, as expected. However, the diagnostic performance of the LDT was adequate as characterized by a sensitivity and specificity of 80 and 64 %, respectively. CONCLUSIONS: This study, designed as a proof of principle, indicated that the LDT is an accurate, robust and automated assay, which adequately and reliably identifies patients presenting with small fiber neuropathy, and therefore has potential for use in large scale clinical studies.


Sujet(s)
Neuropathie des petites fibres/diagnostic , Ubiquitin thiolesterase/métabolisme , Adulte , Technique d'immunofluorescence , Humains , Adulte d'âge moyen , Biais de l'observateur , Neuropathie des petites fibres/métabolisme
16.
Neurobiol Dis ; 73: 83-95, 2015 Jan.
Article de Anglais | MEDLINE | ID: mdl-25220759

RÉSUMÉ

Neurofibrillary tangles composed of hyperphosphorylated fibrillized tau are found in numerous tauopathies including Alzheimer's disease. Increasing evidence suggests that tau pathology can be transmitted from cell-to-cell; however the mechanisms involved in the initiation of tau fibrillization and spreading of disease linked to progression of tau pathology are poorly understood. We show here that intracerebral injections of preformed synthetic tau fibrils into the hippocampus or frontal cortex of young tau transgenic mice expressing mutant human P301L tau induces tau hyperphosphorylation and aggregation around the site of injection, as well as a time-dependent propagation of tau pathology to interconnected brain areas distant from the injection site. Furthermore, we show that the tau pathology as a consequence of injection of tau preformed fibrils into the hippocampus induces selective loss of CA1 neurons. Together, our data confirm previous studies on the seeded induction and the spreading of tau pathology in a different tau transgenic mouse model and reveals neuronal loss associated with seeded tau pathology in tau transgenic mouse brain. These results further validate the utility of the tau seeding model in studying disease transmission, and provide a more complete in vivo tauopathy model with associated neurodegeneration which can be used to investigate the mechanisms involved in tau aggregation and spreading, as well as aid in the search for disease modifying treatments for Alzheimer's disease and related tauopathies.


Sujet(s)
Tauopathies , Protéines tau/administration et posologie , Protéines tau/génétique , Facteurs âges , Analyse de variance , Animaux , Modèles animaux de maladie humaine , Évolution de la maladie , Latéralité fonctionnelle , Régulation de l'expression des gènes/effets des médicaments et des substances chimiques , Régulation de l'expression des gènes/génétique , Hippocampe/effets des médicaments et des substances chimiques , Hippocampe/métabolisme , Hippocampe/anatomopathologie , Humains , Souris , Souris transgéniques , Mutation/génétique , Enchevêtrements neurofibrillaires/métabolisme , Tauopathies/induit chimiquement , Tauopathies/génétique , Tauopathies/anatomopathologie , Protéines tau/composition chimique
17.
Behav Brain Res ; 245: 13-21, 2013 May 15.
Article de Anglais | MEDLINE | ID: mdl-23396167

RÉSUMÉ

The excitatory neurotransmitter l-glutamate is transported into synaptic vesicles by vesicular glutamate transporters (VGluTs) to transmit glutamatergic signals. Changes in their expression have been linked to various brain disorders including schizophrenia, Parkinson's, and Alzheimer's disease. Deleting either the VGluT1 or VGluT2 gene leads to profound developmental and neurological complications and early death, but mice heterozygous for VGluT1 or VGluT2 are viable and thrive. Acquisition, retention and extinction of conditioned visuospatial and emotional responses were compared between VGluT1(+/-) and VGluT2(+/-) mice, and their wildtype littermates, using different water maze procedures, appetitive scheduled conditioning, and conditioned fear protocols. The distinct brain expression profiles of the VGluT1 and -2 isoforms particularly in telencephalic structures, such as neocortex, hippocampus and striatum, are reflected in very specific behavioral changes. VGluT2(+/-) mice were unimpaired in spatial learning tasks and fear extinction. Conversely, VGluT1(+/-) mice displayed spatial extinction learning deficits and markedly impaired fear extinction. These data indicate that VGluT1, but not VGluT2, plays a role in the neural processes underlying inhibitory learning.


Sujet(s)
Extinction (psychologie)/physiologie , Perception de l'espace/physiologie , Transporteur vésiculaire-1 du glutamate/génétique , Transporteur vésiculaire-2 du glutamate/génétique , Animaux , Appétit , Apprentissage par évitement/physiologie , Comportement animal/physiologie , Chimie du cerveau/génétique , Émotions/physiologie , Immunohistochimie , Apprentissage/physiologie , Apprentissage du labyrinthe/physiologie , Mémoire à court terme/physiologie , Souris , Souris knockout , Programme de renforcement , Transporteur vésiculaire-1 du glutamate/déficit , Transporteur vésiculaire-2 du glutamate/déficit
18.
J Histochem Cytochem ; 61(3): 218-35, 2013 Mar.
Article de Anglais | MEDLINE | ID: mdl-23172796

RÉSUMÉ

Although the kidney generally has been regarded as an excellent source of carboxypeptidase M (CPM), little is known about its renal-specific expression level and distribution. This study provides a detailed localization of CPM in healthy and diseased human kidneys. The results indicate a broad distribution of CPM along the renal tubular structures in the healthy kidney. CPM was identified at the parietal epithelium beneath the Bowman's basement membrane and in glomerular mesangial cells. Capillaries, podocytes, and most interstitial cells were CPM negative. Tumor cells of renal cell carcinoma subtypes lose CPM expression upon dedifferentiation. Tissue microarray analysis demonstrated a correlation between low CPM expression and tumor cell type. CPM staining was intense on phagocytotic tumor-associated macrophages. Immunoreactive CPM was also detected in the tumor-associated vasculature. The absence of CPM in normal renal blood vessels points toward a role for CPM in angiogenesis. Coexistence of CPM and the epidermal growth factor receptor (EGFR) was detected in papillary renal cell carcinoma. However, the different subcellular localization of CPM and EGFR argues against an interaction between these h proteins. The description of the distribution of CPM in human kidney forms the foundation for further study of the (patho)physiological activities of CPM in the kidney.


Sujet(s)
Néphrocarcinome/enzymologie , Tumeurs du rein/enzymologie , Rein/enzymologie , Rein/anatomopathologie , Macrophages/enzymologie , Metalloendopeptidases/analyse , Adulte , Néphrocarcinome/vascularisation , Néphrocarcinome/anatomopathologie , Carcinome épidermoïde/enzymologie , Carcinome épidermoïde/anatomopathologie , Récepteurs ErbB/analyse , Protéines liées au GPI/analyse , Humains , Immunohistochimie , Rein/vascularisation , Tumeurs du rein/vascularisation , Tumeurs du rein/anatomopathologie , Macrophages/anatomopathologie , Adulte d'âge moyen , Néovascularisation pathologique/enzymologie , Néovascularisation pathologique/anatomopathologie , Analyse sur puce à tissus
19.
J Neurol ; 258(10): 1852-64, 2011 Oct.
Article de Anglais | MEDLINE | ID: mdl-21472496

RÉSUMÉ

This study aimed at evaluating the performance of a battery of morphological and functional tests for the assessment of small nerve fiber loss in asymptomatic diabetic neuropathy (DNP). Patients diagnosed for ≥10 years with type 1 (n = 10) or type 2 (n = 13) diabetes mellitus (DM) without conventional symptoms or signs of DNP were recruited and compared with healthy controls (n = 18) and patients with overt DNP (n = 5). Intraepidermal nerve fiber density (IENFd) was measured with PGP9.5 immunostaining on punch skin biopsies performed at the distal leg. Functional tests consisted of quantitative sensory testing (QST) for light-touch, cool, warm and heat pain detection thresholds and brain-evoked potentials with electrical (SEPs) and CO(2) laser stimulation [laser-evoked potentials (LEPs)] of hand dorsum and distal leg using small (0.8 mm(2)) and large (20 mm(2)) beam sizes. Results confirmed a state of asymptomatic DNP in DM, but only at the distal leg. Defining a critical small fiber loss as a reduction of IENFd ≤-2 z scores of healthy controls, this state prevailed in type 2 (30%) over type 1 DM (10%) patients despite similar disease duration and current glycemic control. LEPs with the small laser beam performed best in terms of sensitivity (91%), specificity (83%) and area-under-the ROC curve (0.924). Although this performance was not statically different from that of warm and cold detection threshold, LEPs offer an advantage over QST given that they bypass the subjective report and are therefore unbiased by perceptual factors.


Sujet(s)
Biopsie , Neuropathies diabétiques/diagnostic , Potentiels évoqués somatosensoriels , Seuils sensoriels , Adulte , Diabète/anatomopathologie , Diabète/physiopathologie , Neuropathies diabétiques/anatomopathologie , Neuropathies diabétiques/physiopathologie , Diagnostic précoce , Stimulation électrique , Potentiels évoqués somatosensoriels/physiologie , Femelle , Humains , Immunohistochimie , Lasers , Mâle , Adulte d'âge moyen , Neurofibres/anatomopathologie , Sensibilité et spécificité , Seuils sensoriels/physiologie , Peau/innervation
20.
Clin Neurophysiol ; 121(8): 1256-66, 2010 Aug.
Article de Anglais | MEDLINE | ID: mdl-20347388

RÉSUMÉ

OBJECTIVE: To assess the temporal relationship between skin nerve denervation and regeneration (dermal and intra-epidermal fibres, IENF) and functional changes (CO(2) laser-evoked potentials, LEPs, and quantitative sensory tests, QST) after topical cutaneous application of capsaicin. METHODS: Capsaicin (0.075%) was applied to the lateral calf for three consecutive days. QST, LEPs and skin biopsies were performed at baseline and time intervals up to 54days post-capsaicin treatment. Biopsies were immunostained with antibodies for PGP9.5, TRPV1, and GAP-43 (marker of regenerating nerve fibres), and analyzed for IENFs and dermal innervation (for GAP-43). RESULTS: At 1day post-capsaicin, cutaneous thermal sensitivity was reduced, as were LEPs. PGP9.5, TRPV1, and GAP-43 immunoreactive-nerve fibres were almost completely absent. By Day 12, LEPs had fully recovered, but PGP9.5 and TRPV1 IENF continued to be significantly decreased 54days post-capsaicin. In contrast, dermal GAP-43 immunoreactivity closely matched recovery of LEPs. CONCLUSIONS: A good correlation was observed between LEPs and GAP-43 staining, in contrast to PGP9.5 and TRPV1. Laser stimulation is a non-invasive and sensitive method for assessing the initial IENF loss, and regenerating nerve fibres. SIGNIFICANCE: Assessing skin biopsies by PGP9.5 immunostaining alone may miss significant diagnostic and prognostic information regarding regenerating nerve fibres, if other approaches are neglected, e.g. LEPs or GAP-43 immunostaining.


Sujet(s)
Capsaïcine/pharmacologie , Potentiels évoqués somatosensoriels/physiologie , Dégénérescence nerveuse/physiopathologie , Neurofibres/métabolisme , Peau/innervation , Peau/métabolisme , Adulte , Analyse de variance , Femelle , Humains , Immunohistochimie , Lasers à gaz , Mâle , Adulte d'âge moyen , Dégénérescence nerveuse/induit chimiquement , Neurofibres/physiologie , Régénération nerveuse/physiologie , Stimulation physique , Peau/effets des médicaments et des substances chimiques , Peau/physiopathologie , Canaux cationiques TRPV/métabolisme , Facteurs temps
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