Pierre Eid studied biochemistry at the Louis Pasteur University of Strasbourg (France), where he got his Ph.D. in 1978. After a two years postdoc at the Karolinska Institute (Stockholm, Sweden) and at the Institut Pasteur (Paris, France), he worked until 2006 at the “Institut de Recherche Scientifique sur le Cancer” (Villejuif, France) on Type I Interferons, receptors and their signaling pathways. Since 2007 he joined the B. Azzarone’s group on Interleukin-15 in the renal physiopathology at the Inserm 1014 and U1197 units, Paul Brousse Hospital (Villejuif, France). His current interest is centered around the study of Interleukin-15 and the tumoral microenvironment in renal stem cells and renal tumor biomarkers. Address: Villejuif, Île-de-France, France
Supplementary Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15... more Supplementary Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Figure 2 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 2 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Legends for Figures 1-4, Table 1 from Human Renal Cancer Cells Express a Novel Memb... more Supplementary Legends for Figures 1-4, Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Publisher Summary This chapter describes the extraction of alpha interferon (IFN)-receptor comple... more Publisher Summary This chapter describes the extraction of alpha interferon (IFN)-receptor complexes with digitonin. The aim is to obtain an extract of the cellular receptor, stable enough to retain bound ligand and in a form suitable for investigating intrinsic enzyme activities. Certain aspects of the binding of IFN-αs to cellular receptors suggested an interaction related to the growth of the cells. For this reason, exponentially growing cells are taken as starting material. Initial experiments suggest that of the commonly used nonionic detergents, only digitonin serve the purpose. The method described has been developed for 125 I-labeled IFN-α2 bound to cells of the Burkitt line, Daudi; but the method works equally well with other IFN-αs and with other human cells of lymphoid origin. The method described provides an in vitro preparation of a human IFN-α-receptor complex suitable for further biochemical work. The original purpose is to correlate receptor function with biological activity. This inevitably places certain limits on the method. Chromatographic separation of the complex depends upon a well-prepared column. If the gel is too loosely packed, separation is inadequate on a 20 cm column. Columns are washed well with a solution of bovine serum albumin in digitonin prior to use.
Interferons (IFNs) in common with other cytokines activate Janus tyrosine kinases and latent STAT... more Interferons (IFNs) in common with other cytokines activate Janus tyrosine kinases and latent STAT transcription factors upon binding to their cell surface receptor. Type I IFNs bind to a receptor composed of two transmembrane polypeptides, IFNAR1 and IFNAR2, which belong to the class II cytokine receptor family that also includes the cellular receptors for IFN-gamma, interleukin-10 and coagulation protease factor VII (tissue factor). The extracellular domain of the type I IFN receptor chain IFNAR1, has four fibronectin type-III sub-domains. Human IFNAR1 has intrinsic weak affinity for type I IFNs and plays an essential role in transmembrane signaling, formation of a high affinity complex with IFN and the modulation of ligand specificity. In order to characterise the ligand binding site on IFNAR1 we analysed the epitope recognized by the anti-IFNAR1 mAb, 64G12, which inhibits the binding and biological activities of both IFN-alpha and IFN-beta. The target peptide recognized by the 64...
We describe the extraction and stabilization in vitro of discrete complexes of interferon and cel... more We describe the extraction and stabilization in vitro of discrete complexes of interferon and cellular receptor proteins. A homogeneous complex of M r 230000 was extracted at the time of peak receptor binding (30 min). Complex formation was specific for human interferon. At later times a second complex could also be extracted suggesting transfer of interferon to a second site.
Supplementary Figure 4 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 4 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Figure 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Although interleukin-15 (IL-15) is a powerful immunomodulatory factor that has been proposed for ... more Although interleukin-15 (IL-15) is a powerful immunomodulatory factor that has been proposed for cancer immunotherapy, its intratumoral expression may be correlated with tumor progression and/or poor clinical outcome. Therefore, neoplasias potentially sensitive to immunotherapy should be checked for their IL-15 expression and function before choosing immunotherapy protocols. Primary human renal cancer cells (RCC) express a novel form of membrane-bound IL-15 (mb-IL-15), which displays three major original properties: (a) It is expressed as a functional membrane homodimer of 27 kDa, (b) it is shed in the extracellular environment by the metalloproteases ADAM17 and ADAM10, and (c) its stimulation by soluble IL-15 receptor α (s-IL-15Rα) chain triggers a complex reverse signal (mitogen-activated protein kinases, FAK, pMLC) necessary and sufficient to ~induce epithelial-mesenchymal transdifferentiation (EMT), a crucial process in tumor progression whose induction is unprecedented for IL-1...
1. Neuraminidase was obtained by (A) bromelain solubilization or (B) by treatment with N-lauroyls... more 1. Neuraminidase was obtained by (A) bromelain solubilization or (B) by treatment with N-lauroylsarcosine. 2. 5-N-acetyl-2-O-(3-methoxyphenyl)-alpha-D-neuraminic acid, employed as substrate, avoids the interference produced by the thiobarbituric acid method, and is not interfered by the ampholytes. 3. Only about 20% of original enzyme activity was lost after electrofocusing. The sample from procedure A showed two peaks, corresponding to pIs 4.4 and 5.6. The sample from procedure B, having a higher activity, showed only one peak at pI 4.4. 4. Samples A and B showed different Km and hydrolysis rate with N-acetylneuraminyl-lactose and glycophorin A. It was not found significantly different with other substrates: alpha 1-acid glycoprotein, brain gangliosides, 5-N-acetyl-2-O-(3-methoxyphenyl)-alpha-D-neuraminic acid and 2'-(4-methyl umbelliferyl)-alpha-D-N-acetylneuraminic acid.
Renal tubulointerstitial fibrosis is the final common pathway in end-stage renal disease and is c... more Renal tubulointerstitial fibrosis is the final common pathway in end-stage renal disease and is characterized by aberrant accumulation of extracellular matrix (ECM) components secreted by myofibroblasts. Tubular type 2 EMT, induced by TGF-β, plays an important role in renal fibrosis, by participating directly or indirectly in myofibroblasts generation. TGF-β1-induced apoptosis and fibrosis in experimental chronic murine kidney diseases are concomitantly associated with an intrarenal decreased expression of the IL-15 survival factor. Since IL-15 counteracts TGF-β1 effects in different cell models, we analyzed whether (1) human chronic inflammatory nephropathies evolving towards fibrosis could be also characterized by a weak intrarenal IL-15 expression and (2) IL-15 could inhibit epithelial-mesenchymal transition (EMT) and excess matrix deposition in human renal proximal tubular epithelial cells (RPTEC). Our data show that different human chronic kidney diseases are characterized by a...
Supplementary Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15... more Supplementary Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Figure 2 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 2 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Legends for Figures 1-4, Table 1 from Human Renal Cancer Cells Express a Novel Memb... more Supplementary Legends for Figures 1-4, Table 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Publisher Summary This chapter describes the extraction of alpha interferon (IFN)-receptor comple... more Publisher Summary This chapter describes the extraction of alpha interferon (IFN)-receptor complexes with digitonin. The aim is to obtain an extract of the cellular receptor, stable enough to retain bound ligand and in a form suitable for investigating intrinsic enzyme activities. Certain aspects of the binding of IFN-αs to cellular receptors suggested an interaction related to the growth of the cells. For this reason, exponentially growing cells are taken as starting material. Initial experiments suggest that of the commonly used nonionic detergents, only digitonin serve the purpose. The method described has been developed for 125 I-labeled IFN-α2 bound to cells of the Burkitt line, Daudi; but the method works equally well with other IFN-αs and with other human cells of lymphoid origin. The method described provides an in vitro preparation of a human IFN-α-receptor complex suitable for further biochemical work. The original purpose is to correlate receptor function with biological activity. This inevitably places certain limits on the method. Chromatographic separation of the complex depends upon a well-prepared column. If the gel is too loosely packed, separation is inadequate on a 20 cm column. Columns are washed well with a solution of bovine serum albumin in digitonin prior to use.
Interferons (IFNs) in common with other cytokines activate Janus tyrosine kinases and latent STAT... more Interferons (IFNs) in common with other cytokines activate Janus tyrosine kinases and latent STAT transcription factors upon binding to their cell surface receptor. Type I IFNs bind to a receptor composed of two transmembrane polypeptides, IFNAR1 and IFNAR2, which belong to the class II cytokine receptor family that also includes the cellular receptors for IFN-gamma, interleukin-10 and coagulation protease factor VII (tissue factor). The extracellular domain of the type I IFN receptor chain IFNAR1, has four fibronectin type-III sub-domains. Human IFNAR1 has intrinsic weak affinity for type I IFNs and plays an essential role in transmembrane signaling, formation of a high affinity complex with IFN and the modulation of ligand specificity. In order to characterise the ligand binding site on IFNAR1 we analysed the epitope recognized by the anti-IFNAR1 mAb, 64G12, which inhibits the binding and biological activities of both IFN-alpha and IFN-beta. The target peptide recognized by the 64...
We describe the extraction and stabilization in vitro of discrete complexes of interferon and cel... more We describe the extraction and stabilization in vitro of discrete complexes of interferon and cellular receptor proteins. A homogeneous complex of M r 230000 was extracted at the time of peak receptor binding (30 min). Complex formation was specific for human interferon. At later times a second complex could also be extracted suggesting transfer of interferon to a second site.
Supplementary Figure 4 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 4 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Supplementary Figure 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-1... more Supplementary Figure 1 from Human Renal Cancer Cells Express a Novel Membrane-Bound Interleukin-15 that Induces, in Response to the Soluble Interleukin-15 Receptor α Chain, Epithelial-to-Mesenchymal Transition
Although interleukin-15 (IL-15) is a powerful immunomodulatory factor that has been proposed for ... more Although interleukin-15 (IL-15) is a powerful immunomodulatory factor that has been proposed for cancer immunotherapy, its intratumoral expression may be correlated with tumor progression and/or poor clinical outcome. Therefore, neoplasias potentially sensitive to immunotherapy should be checked for their IL-15 expression and function before choosing immunotherapy protocols. Primary human renal cancer cells (RCC) express a novel form of membrane-bound IL-15 (mb-IL-15), which displays three major original properties: (a) It is expressed as a functional membrane homodimer of 27 kDa, (b) it is shed in the extracellular environment by the metalloproteases ADAM17 and ADAM10, and (c) its stimulation by soluble IL-15 receptor α (s-IL-15Rα) chain triggers a complex reverse signal (mitogen-activated protein kinases, FAK, pMLC) necessary and sufficient to ~induce epithelial-mesenchymal transdifferentiation (EMT), a crucial process in tumor progression whose induction is unprecedented for IL-1...
1. Neuraminidase was obtained by (A) bromelain solubilization or (B) by treatment with N-lauroyls... more 1. Neuraminidase was obtained by (A) bromelain solubilization or (B) by treatment with N-lauroylsarcosine. 2. 5-N-acetyl-2-O-(3-methoxyphenyl)-alpha-D-neuraminic acid, employed as substrate, avoids the interference produced by the thiobarbituric acid method, and is not interfered by the ampholytes. 3. Only about 20% of original enzyme activity was lost after electrofocusing. The sample from procedure A showed two peaks, corresponding to pIs 4.4 and 5.6. The sample from procedure B, having a higher activity, showed only one peak at pI 4.4. 4. Samples A and B showed different Km and hydrolysis rate with N-acetylneuraminyl-lactose and glycophorin A. It was not found significantly different with other substrates: alpha 1-acid glycoprotein, brain gangliosides, 5-N-acetyl-2-O-(3-methoxyphenyl)-alpha-D-neuraminic acid and 2'-(4-methyl umbelliferyl)-alpha-D-N-acetylneuraminic acid.
Renal tubulointerstitial fibrosis is the final common pathway in end-stage renal disease and is c... more Renal tubulointerstitial fibrosis is the final common pathway in end-stage renal disease and is characterized by aberrant accumulation of extracellular matrix (ECM) components secreted by myofibroblasts. Tubular type 2 EMT, induced by TGF-β, plays an important role in renal fibrosis, by participating directly or indirectly in myofibroblasts generation. TGF-β1-induced apoptosis and fibrosis in experimental chronic murine kidney diseases are concomitantly associated with an intrarenal decreased expression of the IL-15 survival factor. Since IL-15 counteracts TGF-β1 effects in different cell models, we analyzed whether (1) human chronic inflammatory nephropathies evolving towards fibrosis could be also characterized by a weak intrarenal IL-15 expression and (2) IL-15 could inhibit epithelial-mesenchymal transition (EMT) and excess matrix deposition in human renal proximal tubular epithelial cells (RPTEC). Our data show that different human chronic kidney diseases are characterized by a...
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