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Le Roux, Sandrine
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Publications (4 of 4) Show all publications
Spang, C., Backman, L. J., Le Roux, S., Chen, J. & Danielson, P. (2017). Glutamate signaling through the NMDA receptor reduces the expression of scleraxis in plantaris tendon derived cells. BMC Musculoskeletal Disorders, 18, Article ID 218.
Open this publication in new window or tab >>Glutamate signaling through the NMDA receptor reduces the expression of scleraxis in plantaris tendon derived cells
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2017 (English)In: BMC Musculoskeletal Disorders, ISSN 1471-2474, E-ISSN 1471-2474, Vol. 18, article id 218Article in journal (Refereed) Published
Abstract [en]

Background: A body of evidence demonstrating changes to the glutaminergic system in tendinopathy has recently emerged. This hypothesis was further tested by studying the effects of glutamate on the tenocyte phenotype, and the impact of loading and exposure to glucocorticoids on the glutamate signaling machinery.

Methods: Plantaris tendon tissue and cultured plantaris tendon derived cells were immunohisto-/cytochemically stained for glutamate, N-Methyl-D-Aspartate receptor 1 (NMDAR1) and vesicular glutamate transporter 2 (VGluT2). Primary cells were exposed to glutamate or receptor agonist NMDA. Cell death/viability was measured via LDH/MTS assays, and Western blot for cleaved caspase 3 (c-caspase 3) and cleaved poly (ADP-ribose) polymerase (c-PARP). Scleraxis mRNA (Scx)/protein(SCX) were analyzed by qPCR and Western blot, respectively. A FlexCell system was used to apply cyclic strain. The effect of glucocorticoids was studies by adding dexamethasone (Dex). The mRNA of the glutamate synthesizing enzymes Got1 and Gls, and NMDAR1 protein were measured. Levels of free glutamate were determined by a colorimetric assay.

Results: Immunoreactions for glutamate, VGluT2, and NMDAR1 were found in tenocytes and peritendinous cells in tissue sections and in cultured cells. Cell death was induced by high concentrations of glutamate but not by NMDA. Scleraxis mRNA/protein was down-regulated in response to NMDA/glutamate stimulation. Cyclic strain increased, and Dex decreased, Gls and Got1 mRNA expression. Free glutamate levels were lower after Dex exposure.

Conclusions: In conclusion, NMDA receptor stimulation leads to a reduction of scleraxis expression that may be involved in a change of phenotype in tendon cells. Glutamate synthesis is increased in tendon cells in response to strain and decreased by glucocorticoid stimulation. This implies that locally produced glutamate could be involved in the tissue changes observed in tendinopathy.

Keywords
Glutamate, NMDAR1, Plantaris tendon, Tendinopathy, Scleraxis
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-136330 (URN)10.1186/s12891-017-1575-4 (DOI)000402332800006 ()28545490 (PubMedID)
Available from: 2017-06-21 Created: 2017-06-21 Last updated: 2019-02-22Bibliographically approved
Sloniecka, M., Le Roux, S., Zhou, Q. & Danielson, P. (2016). Substance P Enhances Keratocyte Migration and Neutrophil Recruitment through Interleukin-8. Molecular Pharmacology, 89(2), 215-225
Open this publication in new window or tab >>Substance P Enhances Keratocyte Migration and Neutrophil Recruitment through Interleukin-8
2016 (English)In: Molecular Pharmacology, ISSN 0026-895X, E-ISSN 1521-0111, Vol. 89, no 2, p. 215-225Article in journal (Refereed) Published
Abstract [en]

Keratocytes, the resident cells of the corneal stroma, are responsible for maintaining turnover of this tissue by synthesizing extracellular matrix components. When the cornea is injured, the keratocytes migrate to the wounded site and participate in the stromal wound healing. The neuropeptide substance P (SP), which is also known to be produced by non-neuronal cells, has previously been implicated in epithelial wound healing after corneal injury. Corneal scarring, which occurs in the stroma when the process of wound healing has malfunctioned, is one of the major causes of preventable blindness. This study aimed to elucidate the potential role of SP in keratocyte migration and therefore in stromal wound healing. We report that the expression and secretion of SP in human keratocytes are increased in response to injury in vitro. Moreover, SP enhances the migration of keratocytes by inducing the actin cytoskeleton reorganization and focal adhesion formation through the activation of the phosphatidylinositide 3-kinase and Ras-related C3 botulinum toxin substrate 1/Ras homolog gene family, member A pathway. Furthermore, SP stimulation leads to upregulated expression of the proinflammatory and chemotactic cytokine interleukin-8 (IL-8), which also contributes significantly to SP-enhanced keratocyte migration and is able to attract neutrophils. In addition, the preferred SP receptor, the neurokinin-1 receptor, is necessary to induce keratocyte migration and IL-8 secretion. In conclusion, we describe new mechanisms by which SP enhances migration of keratocytes and recruits neutrophils, two necessary steps in the corneal wound-healing process, which are also likely to occur in other tissue injuries.

National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-117400 (URN)10.1124/mol.115.101014 (DOI)000369305700001 ()26646648 (PubMedID)
Available from: 2016-04-05 Created: 2016-02-29 Last updated: 2018-06-07Bibliographically approved
Roux, S. L., Borbely, G., Słoniecka, M., Backman, L. J. & Danielson, P. (2016). Transforming Growth Factor Beta 1 Modulates the Functional Expression of the Neurokinin-1 Receptor in Human Keratocytes. Current Eye Research, 41(8), 1035-1043
Open this publication in new window or tab >>Transforming Growth Factor Beta 1 Modulates the Functional Expression of the Neurokinin-1 Receptor in Human Keratocytes
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2016 (English)In: Current Eye Research, ISSN 0271-3683, E-ISSN 1460-2202, Vol. 41, no 8, p. 1035-1043Article in journal (Refereed) Published
Abstract [en]

PURPOSE: Transforming growth factor beta 1 (TGF-β1) is a cytokine involved in a variety of processes, such as differentiation of fibroblasts into myofibroblasts. TGF-β1 has also been shown to delay the internalization of the neurokinin-1 receptor (NK-1 R) after its activation by its ligand, the neuropeptide substance P (SP). NK-1 R comprises two naturally occurring variants, a full-length and a truncated form, triggering different cellular responses. SP has been shown to affect important events in the cornea - such as stimulating epithelial cell proliferation - processes that are involved in corneal wound healing and thus in maintaining the transparency of the corneal stroma. An impaired signaling through NK-1 R could thus impact the visual quality. We hypothesize that TGF-β1 modulates the expression pattern of NK-1 R in human corneal stroma cells, keratocytes. The purpose of this study was to test that hypothesis.

METHODS: Cultures of primary keratocytes were set up with cells derived from healthy human corneas, obtained from donated transplantation graft leftovers, and characterized by immunocytochemistry and Western blot. Immunocytochemistry for TGF-β receptors and NK-1 R was performed. Gene expression was assessed with real-time polymerase chain reaction (qPCR).

RESULTS: Expression of TGF-β receptors was confirmed in keratocytes in vitro. Treating the cells with TGF-β1 significantly reduced the gene expression of NK-1 R. Furthermore, immunocytochemistry for NK-1 R demonstrated that it is specifically the expression of the full-length isotype of the receptor that is reduced after treatment with TGF-β1, which was also confirmed with qPCR using a specific probe for the full-length receptor.

CONCLUSIONS: TGF-β1 down-regulates the gene expression of the full-length variant of NK-1 R in human keratocytes, which might impact its signaling pathway and thus explain the known delay in internalization after activation by SP seen with TGF-β1 treatment.

Keywords
Cornea, cytokines, neuropeptides, stroma, substance P
National Category
Basic Medicine
Research subject
Human Anatomy
Identifiers
urn:nbn:se:umu:diva-117110 (URN)10.3109/02713683.2015.1088954 (DOI)000382771600005 ()26673553 (PubMedID)
Available from: 2016-02-22 Created: 2016-02-22 Last updated: 2019-02-22Bibliographically approved
Sloniecka, M., Le Roux, S., Boman, P., Byström, B., Zhou, Q. & Danielson, P. (2015). Expression Profiles of Neuropeptides, Neurotransmitters, and Their Receptors in Human Keratocytes In Vitro and In Situ. PLoS ONE, 10(7), Article ID e0134157.
Open this publication in new window or tab >>Expression Profiles of Neuropeptides, Neurotransmitters, and Their Receptors in Human Keratocytes In Vitro and In Situ
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2015 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 7, article id e0134157Article in journal (Refereed) Published
Abstract [en]

Keratocytes, the quiescent cells of the corneal stroma, play a crucial role in corneal wound healing. Neuropeptides and neurotransmitters are usually associated with neuronal signaling, but have recently been shown to be produced also by non-neuronal cells and to be involved in many cellular processes. The aim of this study was to assess the endogenous intracellular and secreted levels of the neuropeptides substance P (SP) and neurokinin A (NKA), and of the neurotransmitters acetylcholine (ACh), catecholamines (adrenaline, noradrenaline and dopamine), and glutamate, as well as the expression profiles of their receptors, in human primary keratocytes in vitro and in keratocytes of human corneal tissue sections in situ. Cultured keratocytes expressed genes encoding for SP and NKA, and for catecholamine and glutamate synthesizing enzymes, as well as genes for neuropeptide, adrenergic and ACh (muscarinic) receptors. Keratocytes in culture produced SP, NKA, catecholamines, ACh, and glutamate, and expressed neurokinin-1 and -2 receptors (NK-1R and NK-2R), dopamine receptor D-2, muscarinic ACh receptors, and NDMAR1 glutamate receptor. Human corneal sections expressed SP, NKA, NK-1R, NK-2R, receptor D2, choline acetyl transferase (ChAT), M-3, M4 and M-5 muscarinic ACh receptors, glutamate, and NMDAR1, but not catecholamine synthesizing enzyme or the alpha(1) and beta(2) adrenoreceptors, nor M1 receptor. In addition, expression profiles assumed significant differences between keratocytes from the peripheral cornea as compared to those from the central cornea, as well as differences between keratocytes cultured under various serum concentrations. In conclusion, human keratocytes express an array of neuropeptides and neurotransmitters. The cells furthermore express receptors for neuropeptides/neurotransmitters, which suggests that they are susceptible to stimulation by these substances in the cornea, whether of neuronal or non-neuronal origin. As it has been shown that neuropeptides/neurotransmitters are involved in cell proliferation, migration, and angiogenesis, it is possible that they play a role in corneal wound healing.

Place, publisher, year, edition, pages
Public library science, 2015
National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-107294 (URN)10.1371/journal.pone.0134157 (DOI)000358594300054 ()26214847 (PubMedID)
Available from: 2015-08-27 Created: 2015-08-21 Last updated: 2018-06-07Bibliographically approved
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