Publications by Isabelle Fournier
An integrated diagnosis using molecular features is recommended in the 2016 World Health Organiza... more An integrated diagnosis using molecular features is recommended in the 2016 World Health Organization (WHO) classification. Our aim was to explore non-targeted molecular classification using MALDI mass spectrom-etry imaging (MALDI MSI) associated to microproteomics in order to classify anaplastic glioma by integration of clinical data. We used fresh-frozen tissue sections to perform MALDI MSI of proteins based on their digestion pep-tides after in-situ trypsin digestion of the tissue sections and matrix deposition by micro-spraying. The generated 70 μm spatial resolution image datasets were further processed by individual or global segmentation in order to cluster the tissues according to their molecular protein signature. The clustering gives 3 main distinct groups. Within the tissues the ROIs (regions of interest) defined by these groups were used for microproteomics by micro-extraction of the tryptic peptides after on-tissue enzymatic digestion. More than 2500 proteins including 22 alternative proteins (AltProt) are identified by the Shotgun microproteomics. Statistical analysis on the basis of the label free quantification of the proteins shows a similar classification to the MALDI MSI segmentation into 3 groups. Functional analysis performed on each group reveals sub-networks related to neoplasia for group 1, gli-oma with inflammation for group 2 and neurogenesis for group 3. This demonstrates the interest on these new non-targeted large molecular data combining both MALDI MSI and microproteomics data, for tumor classification. This analysis provides new insights into grade III glioma organization. This specific information could allow a more accurate classification of the biopsies according to the prognosis and the identification of potential new targeted therapeutic options. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann. Matrix-assisted laser desorption/ionization mass spectrometry imaging Microproteomics Biochimica et Biophysica Acta xxx (2016) xxx–xxx j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / b b a p a p Please cite this article as: E. Le Rhun, et al., Evaluation of non-supervised MALDI mass spectrometry imaging combined with microproteomics for glioma grade III classification, Biochim. Biophys. Acta (2016), http://dx.
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Saponins are secondary metabolites that are abundant and diversified in echinoderms. Mass spectro... more Saponins are secondary metabolites that are abundant and diversified in echinoderms. Mass spectrometry is increasingly used not only to identify saponin congeners within animal extracts but also to decipher the structure/biological activity relationships of these molecules by determining their inter-organ and inter-individual variability. The usual method requires extensive purification procedures to prepare saponin extracts compatible with mass spectrometry analysis. Here, we selected the sea star Asterias rubens as a model animal to prove that direct analysis of saponins can be performed on tissue sections. We also demonstrated that carboxymethyl cellulose can be used as an embedding medium to facilitate the cryosectioning procedure. Matrix-assisted laser desorption/ ionization (MALDI) imaging was also revealed to afford interesting data on the distribution of saponin molecules within the tissues. We indeed highlight that saponins are located not only inside the body wall of the animals but also within the mucus layer that probably protects the animal against external aggressions.
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Mass spectrometry (MS)-based microproteomics on localized regions of tissue sections was achieved... more Mass spectrometry (MS)-based microproteomics on localized regions of tissue sections was achieved by direct coupling of liquid microjunction microextraction with a nanoscale liquid chromatography-tandem MS, resulting in the identification of >500 protein groups from a region as small as 250μm in diameter representing only a few hundred of cells. The method was applied on the examination of benign and tumor regions initially defined by imaging mass spectrometry (IMS) analysis of a consecutive high grade serous ovarian tumor tissue section. Results identified the higher abundance of eukaryotic translation initiation factors eIF4A, its isoform eIF4A2, and eIF5A and its isoform eIF5A2, and lower abundance of actin-binding proteins OBSCN, TAGLN and CNN3 on tumor regions, concomitant with previous findings. This demonstrates the use of the method for downstream characterization of distinct regions identified by IMS.
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Tissue microenvironment characterization presents a challenge for a better understanding of the f... more Tissue microenvironment characterization presents a challenge for a better understanding of the full complexity of a pathology. Unfortunately, making a precise " picture " of the disease needs an efficient microsampling method coupled to an accurate localization for performing region-dependent proteomics. Here, we present a method that enables rapid and reproducible extraction of proteins from a tissue section to analyze a specific region at a millimeter scale. The method used a liquid-microjunction extraction with conventional detergent solution for proteomics analysis. We successfully performed immunoblotting experiments and showed the possibility to retrieve and identify more than 1400 proteins from a 1-mm diameter spot size on tissue sections with a high degree of reproducibility both qualitatively and quantitatively. Moreover, the small size of the extracted region achieved by this sampling method allows the possibility to perform multiple extractions on different tissue section points. Ten points on a sagittal rat brain tissue section were analyzed and the measured proteins clearly distinguished the different parts of the brain, thus permitting precise functional mapping. We thus demonstrate that with this technology, it is possible to map the tissue microenvironment and gain an understanding of the molecular mechanisms at millimeter resolution.
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Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic... more Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic burden on the public and private sectors worldwide. Although several studies have been conducted to identify the molecular progression of injury sequel due from the lesion site, still the exact underlying mechanisms and pathways of injury development have not been fully elucidated. In this work, based on OMICs, 3D matrix-assisted laser desorption ionization (MALDI) imaging, cytokines arrays, confocal imaging we established for the first time that molecular and cellular processes occurring after SCI are altered between the lesion proximity, i.e. rostral and caudal segments nearby the lesion (R1-C1) whereas segments distant from R1-C1, i.e. R2-C2 and R3-C3 levels coexpressed factors implicated in neurogenesis. Delay in T regulators recruitment between R1 and C1 favor discrepancies between the two segments. This is also reinforced by presence of neurites outgrowth inhibitors in C1, absent in R1. Moreover, the presence of immunoglobulins (IgGs) in neurons at the lesion site at 3 days, validated by mass spectrometry, may present additional factor that contributes to limited regeneration. Treatment in vivo with anti-CD20 one hour after SCI did not improve locomotor function and decrease IgG expression. These results open the door of a novel view of the SCI treatment by considering the C1 as the therapeutic target.
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Hydrogen/deuterium exchange mass spectrometric (H/DXMS) methods for protein structural analysis a... more Hydrogen/deuterium exchange mass spectrometric (H/DXMS) methods for protein structural analysis are conventionally performed in solution. We present Tissue Deuterium Exchange Mass Spectrometry (TDXMS), a method to directly monitor deuterium uptake on tissue, as a means to better approximate the deuterium exchange behavior of proteins in their native microenvironment. Using this method, a difference in deuterium uptake behavior was observed when the same proteins were monitored in solution and on tissue. The higher maximum deuterium uptake at equilibrium for all proteins analyzed in solution suggests a more open conformation in the absence of interacting partners normally observed on tissue. We also demonstrate a difference in the deuterium uptake behavior of a few proteins across different morphological regions of the same tissue section. Modifications of the total number of hydrogens exchanged, as well as the kinetics of exchange, were both observed. These results provide information on the implication of protein interactions with partners as well as on the conformational changes related to these interactions, and illustrate the importance of examining protein deuterium exchange behavior in the presence of its specific microenvironment directly at the level of tissues.
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Here we describe a new instrument (SpiderMass) designed for in vivo and real-time analysis. In th... more Here we describe a new instrument (SpiderMass) designed for in vivo and real-time analysis. In this instrument ion production is performed remotely from the MS instrument and the generated ions are transported in real-time to the MS analyzer. Ion production is promoted by Resonant Infrared Laser Ablation (RIR-LA) based on the highly effective excitation of O-H bonds in water molecules naturally present in most biological samples. The retrieved molecular patterns are specific to the cell phenotypes and benign versus cancer regions of patient biopsies can be easily differentiated. We also demonstrate by analysis of human skin that SpiderMass can be used under in vivo conditions with minimal damage and pain. Furthermore SpiderMass can also be used for real-time drug metabolism and pharmacokinetic (DMPK) analysis or food safety topics. SpiderMass is thus the first MS based system designed for in vivo real-time analysis under minimally invasive conditions. Mass Spectrometry (MS) is one of the analytical techniques with the widest accessible range of applications. Its strength relies in its ability to retrieve molecular information from complex mixtures with the possibility to identify each detected compound, as well as the broad panel of MS-based techniques that allow the design of instruments of various geometries, performances, characteristics and capabilities. MS can be used for applications as remote as analyzing solar wind during space missions 1,2 or trying to get a deeper coverage of the proteome of human cells 3,4. The last decades have witnessed the important changes within the field of MS slowly moving from conventional analysis of extracted samples introduced into the mass spectrometer to the analysis of sample tissue pieces. This has led to the emergence and development of various imaging technologies starting with SIMS MS imaging in the 60's 5,6 to direct MALDI MS analyses 7 and MALDI MS imaging 8. More recently, because imaging techniques are often performed under high vacuum conditions that require a careful preparation of the samples before analysis, many novel ion production methods operating under atmospheric pressure conditions such as AP-MALDI 9,10 , Desorption ElectroSpray Ionization (DESI) 11 or Laser Ablation ElectroSpray Ionization (LAESI) 12 were introduced. The growing number of studies using surface analysis demonstrate the importance of retrieving molecular information and correlate to a location on the surface (spatially resolved analysis). In biological and clinical applications, it has been clearly demonstrated that the spatially resolved molecular information retrieved can be correlated to a physiological or physiopathological event 13–17. Even though enormous progress has been achieved with the possibility to analyze surfaces or sometimes small pieces of material extracted from bigger objects or living organisms, the next challenge for MS is to translate from ex vivo analysis to in vivo conditions at the level of human beings. Being able to get molecular information directly in vivo is an important goal and already exponentially broaden the possibilities of MS. DESI was one of the first technologies used to apply MS in examining living organisms 18. DESI can be applied to search for pesticides from intact fruits or vegetables or by using proper solvents for finding traces of drugs or gunpowder directly from the hands 18–20. Many other technologies have emerged in the past decade 21 , some of them have found applications in daily life such as PaperSpray combined with a miniaturized MS instrument that is implemented at airport security control gates 22,23. The next Holy Grail to be reached for MS is then to be able to perform in vivo analysis of human beings for healthcare
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Numerous applications of ambient Mass Spectrometry (MS) have been demonstrated over the past deca... more Numerous applications of ambient Mass Spectrometry (MS) have been demonstrated over the past decade. They promoted the emergence of various micro-sampling techniques such as Laser Ablation/ Droplet Capture (LADC). LADC consists in the ablation of analytes from a surface and their subsequent capture in a solvent droplet which can then be analyzed by MS. LADC is thus generally performed in the UV or IR range, using a wavelength at which analytes or the matrix absorb. In this work, we explore the potential of visible range LADC (532 nm) as a micro-sampling technology for large-scale proteomics analyses. We demonstrate that biomolecule analyses using 532 nm LADC are possible, despite the low absorbance of biomolecules at this wavelength. This is due to the preponderance of an indirect substrate-mediated ablation mechanism at low laser energy which contrasts with the conventional direct ablation driven by sample absorption. Using our custom LADC system and taking advantage of this substrate-mediated ablation mechanism, we were able to perform large-scale proteomic analyses of micro-sampled tissue sections and demonstrated the possible identification of proteins with relevant biological functions. Consequently, the 532 nm LADC technique offers a new tool for biological and clinical applications. Technological evolution in the field of MS over the past decade has promoted the development of new tools for analyses of crude samples or object surfaces. Accordingly, ambient MS has become a powerful tool for direct analysis of native samples with minimal sample preparation. Consequently, fields of application of ambient MS have widened to human health and environmental studies, as various sensitive objects and organisms such as living tissues or plants can be analyzed using this technique 1. Most of the ambient MS techniques are either derived from Electrospray Ionization (ESI) or from laser based technologies. The former includes Desorption ElectroSpray Ionization (DESI) 2,3 and Easy Ambient Sonic Spray Ionization (EASSI) 4 , both generating ions by exposing the sample to charged droplets, whereas the latter includes Laser Desorption Ionization (LDI) or Laser Ablation (LA). Several concepts have been explored in the field of laser-based ambient MS techniques. Depending on the technique, both desorption and ionization of molecules are either achieved directly from a pristine sample or from a mixture of the sample embedded within a suitable matrix material. Techniques involving a matrix material are derived from Matrix Assisted Laser Desorption/Ionization (MALDI) and include Atmospheric Pressure MALDI (AP-MALDI) 5–7. Desorption and ionization can also be decoupled when molecules are first ejected, and then captured and subsequently ionized. The capture is commonly achieved by a spray of charged solvent droplets that are delivered under ESI conditions, allowing for direct analysis of the sampled material by MS. Techniques using this procedure are Laser Desorption Electrospray Ionization (LDESI) 8,9 , Matrix-Assisted Laser Desorption Electrospray Ionization (MALDESI) 10,11 , Electrospray Laser Desorption Ionization (ELDI) 12 , Laser Electrospray Mass Spectrometry (LEMS) 13,14 or Laser Ablation Electrospray Ionization (LAESI) 15. Alternatively, ejected molecules can be captured into neutral solvent droplets followed by indirect MS analysis for Laser Ablation/Droplet Capture (LADC) 16–19 or be captured by a continuous flow of solvent for direct MS analysis 20,21. These laser-based
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BACKGROUND:
The pathogenesis of acne vulgaris involves several phases including androgen-dependen... more BACKGROUND:
The pathogenesis of acne vulgaris involves several phases including androgen-dependent hyper-seborrhea, colonization by Propionibacterium acnes, and inflammation. Recent investigations have shown that in fact P. acnes provokes the activation of the inflammasome present in macrophages and dendritic cells. This signaling pathway leads to excessive production of interleukin IL-1β, a proinflammatory cytokine. Nevertheless, these well-studied phenomena in acne fail to elucidate the mechanisms responsible for the appearance of different lesions.
METHODS:
We investigate response pathways for specific acne lesions such as microcysts and papules using shot-gun proteomic followed by systemic biology and transcriptomic approaches.
RESULTS:
Results show that most of the proteins identified as differentially expressed between the normal and acne tissue biopsies associated with the immune system response were identified as highly or exclusively expressed in the papule biopsies. They were also expressed in microcysts, but in lower amounts compared to those in papules. These results are supported by the identification of CAMP factor protein produced by P. acnes in microcysts, indicating its enhanced proliferation in this type of lesion CONCLUSIONS: As CAMP factor protein was not detected in papule biopsies, we can see a clear delineation in the stages of progression of acne pathogenesis, which begins with a hyphenated inflammatory response in the papule stage, followed by imbalance of lipid production, which in turn triggers the enhanced proliferation of P. acnes.
GENERAL SIGNIFICANCE:
We demonstrate that expression inflammation varies across the two types of lesions, suggesting different pathways enhanced as a function of the progression of P. acnes.
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Despite being controversial for many years, it is now widely accepted that the Matrix Assisted La... more Despite being controversial for many years, it is now widely accepted that the Matrix Assisted Laser Des-orption/Ionization (MALDI) process leads to the ejection of aggregates or clusters in addition to isolated particles and ions. Clusters composed of analyte surrounded by matrix molecules exhibit a distribution in size and are relatively stable. Several studies have demonstrated that these clusters do not necessarily fully desolvate before ion extraction, but rather progressively along the ion path. This leads to a decrease in analytical performances since these non-desolvated clusters do not contribute to the ion signal but also generate an important chemical background noise. Therefore, proper cluster desolvation before ion extraction in order to release naked analyte ions in the gas phase holds great promise of improved mass spectra sensitivity. This work presents a setup allowing cluster desolvation using nanosecond pulses of a near-infrared laser that intercepts the expanding MALDI plume before ion extraction.
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Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic... more Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic burden on the public and private sectors worldwide. Although several studies have been conducted to identify the molecular progression of injury sequel due from the lesion site, still the exact underlying mechanisms and pathways of injury development have not been fully elucidated. In this work, based on OMICs, 3D matrix-assisted laser desorption ion-ization (MALDI) imaging, cytokines arrays, confocal imaging we established for the first time that molecular and cellular processes occurring after SCI are altered between the lesion proximity, i.e. rostral and caudal segments nearby the lesion (R1-C1) whereas segments distant from R1-C1, i.e. R2-C2 and R3-C3 levels coexpressed factors implicated in neurogenesis. Delay in T regulators recruitment between R1 and C1 favor discrepancies between the two segments. This is also reinforced by presence of neurites outgrowth inhibitors in C1, absent in R1. Moreover, the presence of immunoglobulins (IgGs) in neu-rons at the lesion site at 3 days, validated by mass spec-trometry, may present additional factor that contributes to limited regeneration. Treatment in vivo with anti-CD20 one hour after SCI did not improve locomotor function and decrease IgG expression. These results open the door of a novel view of the SCI treatment by considering the C1 as the therapeutic target. Molecular & Cellular Proteomics 15: 10.1074/mcp.M115.057794, 2641–2670, 2016. Spinal cord injury (SCI) 1 belongs to the serious, currently incurable disorders of the central nervous system (CNS), that are often accompanied by a permanent disability (1). Most SCI are related to traumatic spinal cord damages induced by road trauma, falls, or sport injuries (diving). Among the hallmark features of SCI is the axonal disruption in the spinal cord, which is often caused by fractured intervertebral disc or vertebrate. This primary event is followed by a progressive cascade of secondary deleterious reactions spreading to the adjacent spared tissue leading to a worsening of the neuro-logical status (2, 3). Although axonal regeneration is initiated, it is hampered by a combination of local factors that include severe inflammation, lack of trophic support and development of an inhibitory scar-forming environment. In fact, the regen-erative capacity of the central nervous system is particularly challenged in SCI as multiple cues converge to act as a chemical and physical barrier for the repair process (4, 5). It is now acknowledged that inflammation is one of the major key player that determines abortive axonal repair in SCI. Thus, although the immune response is recognized as primordial to preserve tissue homeostasis, the spatio-temporal course of inflammation in SCI is not favorable to axonal regeneration. Acute inflammation develops hours to days following initial spinal cord trauma and is triggered by the axonal damage and
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Biomaterials capable of controlling the release of multiple growth factors (GFs) could potentiall... more Biomaterials capable of controlling the release of multiple growth factors (GFs) could potentially
promote the integration of co-transplanted neural progenitor cells (NPCs) and stimulate the plasticity
and regenerability of the lesioned spinal cord. As a first step towards the employment of such a
vehicle for cell therapy, this study examined the capability of an alginate–sulphate/alginate scaffold,
able to capture and rigorously control the release of GFs, to promote the expansion and lineage
differentiation of NPCs in vitro. Epidermal growth factor (EGF) and fibroblast growth factor-2 (bFGF)
were affinity-bound to alginate–sulphate (200 ng/scaffold) and the bioconjugates were mixed with
partially calcium-crosslinked alginate. NPCs isolated from 18 day-old rat embryo brains and seeded
into the scaffold during preparation were found to proliferate and differentiate within the vehicle.
A continuous release of both bFGF and EGF was noted for a period of 21 days. The concentrations
of released GFs were sufficient to promote extensive NPC proliferation at initial cultivation times;
the number of neurospheres in the scaffold was twice the number found in the 2D cultures
supplemented with 20 ng/ml each factor every 3 days. Between days 10–14, when the GF concentrations
had substantially declined, extensive cell migration from the neurospheres as well as lineage
differentiation were noted in the scaffold; immunocytochemical analyses confirmed the presence of
neurons, astrocytes and oligodendrocytes.The scaffold has a potential to serve as cell delivery
vehicle, with proven capability to promote cell retention and expansion, while enabling NPC lineage
differentiation in situ.
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Saponins are secondary metabolites that are abundant
and diversified in echinoderms. Mass spectro... more Saponins are secondary metabolites that are abundant
and diversified in echinoderms. Mass spectrometry is
increasingly used not only to identify saponin congeners within
animal extracts but also to decipher the structure/biological
activity relationships of these molecules by determining their
inter-organ and inter-individual variability. The usual method
requires extensive purification procedures to prepare saponin
extracts compatible with mass spectrometry analysis. Here,
we selected the sea star Asterias rubens as a model animal to
prove that direct analysis of saponins can be performed on
tissue sections.We also demonstrated that carboxymethyl cellulose
can be used as an embedding medium to facilitate the
cryosectioning procedure. Matrix-assisted laser desorption/
ionization (MALDI) imaging was also revealed to afford interesting
data on the distribution of saponin molecules within
the tissues. We indeed highlight that saponins are located not
only inside the body wall of the animals but also within the
mucus layer that probably protects the animal against external
aggressions.
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Papers by Isabelle Fournier
Scientific reports, Jan 17, 2015
Numerous applications of ambient Mass Spectrometry (MS) have been demonstrated over the past deca... more Numerous applications of ambient Mass Spectrometry (MS) have been demonstrated over the past decade. They promoted the emergence of various micro-sampling techniques such as Laser Ablation/Droplet Capture (LADC). LADC consists in the ablation of analytes from a surface and their subsequent capture in a solvent droplet which can then be analyzed by MS. LADC is thus generally performed in the UV or IR range, using a wavelength at which analytes or the matrix absorb. In this work, we explore the potential of visible range LADC (532 nm) as a micro-sampling technology for large-scale proteomics analyses. We demonstrate that biomolecule analyses using 532 nm LADC are possible, despite the low absorbance of biomolecules at this wavelength. This is due to the preponderance of an indirect substrate-mediated ablation mechanism at low laser energy which contrasts with the conventional direct ablation driven by sample absorption. Using our custom LADC system and taking advantage of this substra...
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Tissue top-down microproteomics was performed on 3 brain regions, leading to the characterization... more Tissue top-down microproteomics was performed on 3 brain regions, leading to the characterization of 123 reference proteins. Moreover, 8 alternative proteins from alternative open reading frames (AltORF) were identified. Some proteins display specific post-translational modification profiles or truncation linked to the brain regions and their functions. Systems biology analysis performed on the microproteome identified in each region allowed to associate sub-networks with the functional physiology of each brain region. Back correlation of the identified proteins from the microproteome with tissue localization was then performed by MALDI mass spectrometry imaging. As an example, mapping of the distribution of the matrix metallopeptidase 3-cleaved C-terminal fragment of alpha-synuclein (aa 95-140) identified its specific distribution along the hippocampal dentate gyrus. Taken together, we established the molecular physiome of 3 rat brain regions through reference and hidden proteome characterization.
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Journal of Proteomics, 2008
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Current Pharmaceutical Design, 2007
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médecine/sciences, 2007
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Publications by Isabelle Fournier
The pathogenesis of acne vulgaris involves several phases including androgen-dependent hyper-seborrhea, colonization by Propionibacterium acnes, and inflammation. Recent investigations have shown that in fact P. acnes provokes the activation of the inflammasome present in macrophages and dendritic cells. This signaling pathway leads to excessive production of interleukin IL-1β, a proinflammatory cytokine. Nevertheless, these well-studied phenomena in acne fail to elucidate the mechanisms responsible for the appearance of different lesions.
METHODS:
We investigate response pathways for specific acne lesions such as microcysts and papules using shot-gun proteomic followed by systemic biology and transcriptomic approaches.
RESULTS:
Results show that most of the proteins identified as differentially expressed between the normal and acne tissue biopsies associated with the immune system response were identified as highly or exclusively expressed in the papule biopsies. They were also expressed in microcysts, but in lower amounts compared to those in papules. These results are supported by the identification of CAMP factor protein produced by P. acnes in microcysts, indicating its enhanced proliferation in this type of lesion CONCLUSIONS: As CAMP factor protein was not detected in papule biopsies, we can see a clear delineation in the stages of progression of acne pathogenesis, which begins with a hyphenated inflammatory response in the papule stage, followed by imbalance of lipid production, which in turn triggers the enhanced proliferation of P. acnes.
GENERAL SIGNIFICANCE:
We demonstrate that expression inflammation varies across the two types of lesions, suggesting different pathways enhanced as a function of the progression of P. acnes.
promote the integration of co-transplanted neural progenitor cells (NPCs) and stimulate the plasticity
and regenerability of the lesioned spinal cord. As a first step towards the employment of such a
vehicle for cell therapy, this study examined the capability of an alginate–sulphate/alginate scaffold,
able to capture and rigorously control the release of GFs, to promote the expansion and lineage
differentiation of NPCs in vitro. Epidermal growth factor (EGF) and fibroblast growth factor-2 (bFGF)
were affinity-bound to alginate–sulphate (200 ng/scaffold) and the bioconjugates were mixed with
partially calcium-crosslinked alginate. NPCs isolated from 18 day-old rat embryo brains and seeded
into the scaffold during preparation were found to proliferate and differentiate within the vehicle.
A continuous release of both bFGF and EGF was noted for a period of 21 days. The concentrations
of released GFs were sufficient to promote extensive NPC proliferation at initial cultivation times;
the number of neurospheres in the scaffold was twice the number found in the 2D cultures
supplemented with 20 ng/ml each factor every 3 days. Between days 10–14, when the GF concentrations
had substantially declined, extensive cell migration from the neurospheres as well as lineage
differentiation were noted in the scaffold; immunocytochemical analyses confirmed the presence of
neurons, astrocytes and oligodendrocytes.The scaffold has a potential to serve as cell delivery
vehicle, with proven capability to promote cell retention and expansion, while enabling NPC lineage
differentiation in situ.
and diversified in echinoderms. Mass spectrometry is
increasingly used not only to identify saponin congeners within
animal extracts but also to decipher the structure/biological
activity relationships of these molecules by determining their
inter-organ and inter-individual variability. The usual method
requires extensive purification procedures to prepare saponin
extracts compatible with mass spectrometry analysis. Here,
we selected the sea star Asterias rubens as a model animal to
prove that direct analysis of saponins can be performed on
tissue sections.We also demonstrated that carboxymethyl cellulose
can be used as an embedding medium to facilitate the
cryosectioning procedure. Matrix-assisted laser desorption/
ionization (MALDI) imaging was also revealed to afford interesting
data on the distribution of saponin molecules within
the tissues. We indeed highlight that saponins are located not
only inside the body wall of the animals but also within the
mucus layer that probably protects the animal against external
aggressions.
Papers by Isabelle Fournier
The pathogenesis of acne vulgaris involves several phases including androgen-dependent hyper-seborrhea, colonization by Propionibacterium acnes, and inflammation. Recent investigations have shown that in fact P. acnes provokes the activation of the inflammasome present in macrophages and dendritic cells. This signaling pathway leads to excessive production of interleukin IL-1β, a proinflammatory cytokine. Nevertheless, these well-studied phenomena in acne fail to elucidate the mechanisms responsible for the appearance of different lesions.
METHODS:
We investigate response pathways for specific acne lesions such as microcysts and papules using shot-gun proteomic followed by systemic biology and transcriptomic approaches.
RESULTS:
Results show that most of the proteins identified as differentially expressed between the normal and acne tissue biopsies associated with the immune system response were identified as highly or exclusively expressed in the papule biopsies. They were also expressed in microcysts, but in lower amounts compared to those in papules. These results are supported by the identification of CAMP factor protein produced by P. acnes in microcysts, indicating its enhanced proliferation in this type of lesion CONCLUSIONS: As CAMP factor protein was not detected in papule biopsies, we can see a clear delineation in the stages of progression of acne pathogenesis, which begins with a hyphenated inflammatory response in the papule stage, followed by imbalance of lipid production, which in turn triggers the enhanced proliferation of P. acnes.
GENERAL SIGNIFICANCE:
We demonstrate that expression inflammation varies across the two types of lesions, suggesting different pathways enhanced as a function of the progression of P. acnes.
promote the integration of co-transplanted neural progenitor cells (NPCs) and stimulate the plasticity
and regenerability of the lesioned spinal cord. As a first step towards the employment of such a
vehicle for cell therapy, this study examined the capability of an alginate–sulphate/alginate scaffold,
able to capture and rigorously control the release of GFs, to promote the expansion and lineage
differentiation of NPCs in vitro. Epidermal growth factor (EGF) and fibroblast growth factor-2 (bFGF)
were affinity-bound to alginate–sulphate (200 ng/scaffold) and the bioconjugates were mixed with
partially calcium-crosslinked alginate. NPCs isolated from 18 day-old rat embryo brains and seeded
into the scaffold during preparation were found to proliferate and differentiate within the vehicle.
A continuous release of both bFGF and EGF was noted for a period of 21 days. The concentrations
of released GFs were sufficient to promote extensive NPC proliferation at initial cultivation times;
the number of neurospheres in the scaffold was twice the number found in the 2D cultures
supplemented with 20 ng/ml each factor every 3 days. Between days 10–14, when the GF concentrations
had substantially declined, extensive cell migration from the neurospheres as well as lineage
differentiation were noted in the scaffold; immunocytochemical analyses confirmed the presence of
neurons, astrocytes and oligodendrocytes.The scaffold has a potential to serve as cell delivery
vehicle, with proven capability to promote cell retention and expansion, while enabling NPC lineage
differentiation in situ.
and diversified in echinoderms. Mass spectrometry is
increasingly used not only to identify saponin congeners within
animal extracts but also to decipher the structure/biological
activity relationships of these molecules by determining their
inter-organ and inter-individual variability. The usual method
requires extensive purification procedures to prepare saponin
extracts compatible with mass spectrometry analysis. Here,
we selected the sea star Asterias rubens as a model animal to
prove that direct analysis of saponins can be performed on
tissue sections.We also demonstrated that carboxymethyl cellulose
can be used as an embedding medium to facilitate the
cryosectioning procedure. Matrix-assisted laser desorption/
ionization (MALDI) imaging was also revealed to afford interesting
data on the distribution of saponin molecules within
the tissues. We indeed highlight that saponins are located not
only inside the body wall of the animals but also within the
mucus layer that probably protects the animal against external
aggressions.