umu.sePublications
Change search
Link to record
Permanent link

Direct link
BETA
Danielson, Patrik
Publications (10 of 63) Show all publications
Sloniecka, M. & Danielson, P. (2018). Acetylcholine decreases formation of myofibroblasts and excessive extracellular matrix components production in an in vitro human corneal fibrosis model. Paper presented at Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), APR 29-MAY 03, 2018, Honolulu, HI. Investigative Ophthalmology and Visual Science, 59(9)
Open this publication in new window or tab >>Acetylcholine decreases formation of myofibroblasts and excessive extracellular matrix components production in an in vitro human corneal fibrosis model
2018 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 59, no 9Article in journal, Meeting abstract (Other academic) Published
Abstract [en]

Purpose : Acetylcholine (ACh) is a neurotransmitter present in corneal stroma and produced by keratocytes. It has been shown to play a role in processes important for wound healing. Based on literature and our previous studies, we hypothesize that ACh regulates expression of extracellular matrix (ECM) components that are overexpressed during fibrosis, such as collagens, proteoglycans, fibronectin and metalloproteinases, in a protective manner during corneal fibrosis, i.e. decreasing their expression.

Methods : Primary keratocytes were isolated from healthy human corneas obtained from the local cornea bank and grown in presence of 10% fetal bovine serum in order to obtain corneal fibroblasts. A corneal fibrosis in vitro model, in which fibroblasts are stimulated with transforming growth factor beta 1 (TGF-β1) and stable vitamin C, was used throughout this study. Contractile ability of myofibroblasts was tested using a cell contraction assay. Gene expression of ECM components (collagen I, collagen III, collagen V and lumican), markers of fibrosis (α-smooth muscle actin [α-SMA] and fibronectin), and metalloproteinases (MMP2, MMP9 and MMP12) were assessed by qRT-PCR. Intracellular production and secretion of pro-collagen I and lumican was determined by ELISA. α-SMA protein expression was assessed by western blot.

Results : ACh decreased the contractile ability of the newly formed myofibroblasts. ACh significantly decreased gene expression of collagen I, collagen III and collagen V in myofibroblasts. Moreover, ACh treated cells produced and secreted less pro-collagen I. Gene expression of lumican was unaffected by ACh treatment up to day 2 but significantly decreased by day 4. However, no differences in lumican protein level, both intracellular and secreted, were found. ACh downregulated expression of both α-SMA and fibronectin genes. Additionally, α-SMA protein expression was also diminished in ACh treated cells. Furthermore, ACh treatment resulted in downregulation of MMP2, MMP9 and MMP12 genes.

Conclusions : Our results are consistent with our hypothesis that ACh regulates expression of various collagens, lumican, fibronectin and metalloproteinases during corneal fibrosis in vitro, in a way that it diminishes their expression, both on RNA and protein levels. In conclusion, ACh seems to provide protection against formation of fibrosis in human cornea.

Place, publisher, year, edition, pages
The Association for Research in Vision and Ophthalmology, Inc., 2018
National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-152274 (URN)000442912506279 ()
Conference
Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), APR 29-MAY 03, 2018, Honolulu, HI
Funder
Swedish Research Council, 521-2013-2612The Kempe Foundations, JCK-1222Swedish Society of Medicine, SLS-410021Swedish Society of Medicine, SLS-410021Västerbotten County Council, VLL-363161Västerbotten County Council, VLL-549761
Available from: 2018-10-02 Created: 2018-10-02 Last updated: 2018-10-02Bibliographically approved
Chen, J., Zhang, W., Backman, L. J., Kelk, P. & Danielson, P. (2018). Mechanical stress potentiates the differentiation of periodontal ligament stem cells into keratocytes. British Journal of Ophthalmology, 102(4), 562-569
Open this publication in new window or tab >>Mechanical stress potentiates the differentiation of periodontal ligament stem cells into keratocytes
Show others...
2018 (English)In: British Journal of Ophthalmology, ISSN 0007-1161, E-ISSN 1468-2079, Vol. 102, no 4, p. 562-569Article in journal (Refereed) Published
Abstract [en]

Aims To explore the role of corneal-shaped static mechanical strain on the differentiation of human periodontal ligament stem cells (PDLSCs) into keratocytes and the possible synergistic effects of mechanics and inducing medium. Methods PDLSCs were exposed to 3% static dome-shaped mechanical strain in a Flexcell Tension System for 3 days and 7 days. Keratocyte phenotype was determined by gene expression of keratocyte markers. Keratocyte differentiation (inducing) medium was introduced in the Flexcell system, either continuously or intermittently combined with mechanical stimulation. The synergistic effects of mechanics and inducing medium on keratocyte differentiation was evaluated by gene and protein expression of keratocyte markers. Finally, a multilamellar cell sheet was assembled by seeding PDLSCs on a collagen membrane and inducing keratocyte differentiation. The transparency of the cell sheet was assessed, and typical markers of native human corneal stroma were evaluated by immunofluorescence staining. Results Dome-shaped mechanical stimulation promoted PDLSCs to differentiate into keratocytes, as shown by the upregulation of ALDH3A1, CD34, LUM, COL I and COL V. The expression of integrins were also upregulated after mechanical stimulation, including integrin alpha 1, alpha 2, beta 1 and non-muscle myosin II B. A synergistic effect of mechanics and inducing medium was found on keratocyte differentiation. The cell sheets were assembled under the treatment of mechanics and inducing medium simultaneously. The cell sheets were transparent, multilamellar and expressed typical markers of corneal stroma. Conclusion Dome-shaped mechanical stimulation promotes differentiation of PDLSCs into keratocytes and has synergistic effects with inducing medium. Multilamellar cell sheets that resemble native human corneal stroma show potential for future clinical applications.

Place, publisher, year, edition, pages
BMJ Publishing Group Ltd, 2018
Keywords
PDLSCs, corneal stroma, mechanics, inducing medium, differentiation, cell-sheet
National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-147334 (URN)10.1136/bjophthalmol-2017-311150 (DOI)000429732500026 ()29306866 (PubMedID)
Available from: 2018-05-17 Created: 2018-05-17 Last updated: 2019-02-22Bibliographically approved
Sloniecka, M. & Danielson, P. (2018). Substance P Promotes Fibrosis in Human Corneal Stroma. Wound Repair and Regeneration, 26(1), A22-A22, Article ID N4.05.
Open this publication in new window or tab >>Substance P Promotes Fibrosis in Human Corneal Stroma
2018 (English)In: Wound Repair and Regeneration, ISSN 1067-1927, E-ISSN 1524-475X, Vol. 26, no 1, p. A22-A22, article id N4.05Article in journal, Meeting abstract (Other academic) Published
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-148839 (URN)000430308600088 ()
Available from: 2018-06-12 Created: 2018-06-12 Last updated: 2018-06-12Bibliographically approved
Sloniecka, M. U. & Danielson, P. (2017). Acetylcholine regulates expression of lumican and collagen i in keratocytes in quiescent state and after transitioning to fibroblasts and myofibroblasts. Wound Repair and Regeneration, 25(4), A32-A32
Open this publication in new window or tab >>Acetylcholine regulates expression of lumican and collagen i in keratocytes in quiescent state and after transitioning to fibroblasts and myofibroblasts
2017 (English)In: Wound Repair and Regeneration, ISSN 1067-1927, E-ISSN 1524-475X, Vol. 25, no 4, p. A32-A32Article in journal, Meeting abstract (Other academic) Published
Place, publisher, year, edition, pages
WILEY, 2017
National Category
Dermatology and Venereal Diseases
Identifiers
urn:nbn:se:umu:diva-143673 (URN)000416631700136 ()
Available from: 2018-01-16 Created: 2018-01-16 Last updated: 2018-06-09Bibliographically approved
Chen, J., Lan, J., Liu, D., Backman, L. J., Zhang, W., Zhou, Q. & Danielson, P. (2017). Ascorbic Acid Promotes the Stemness of Corneal Epithelial Stem/Progenitor Cells and Accelerates Epithelial Wound Healing in the Cornea. Stem Cells Translational Medicine, 6(5), 1356-1365
Open this publication in new window or tab >>Ascorbic Acid Promotes the Stemness of Corneal Epithelial Stem/Progenitor Cells and Accelerates Epithelial Wound Healing in the Cornea
Show others...
2017 (English)In: Stem Cells Translational Medicine, ISSN 2157-6564, E-ISSN 2157-6580, Vol. 6, no 5, p. 1356-1365Article in journal (Refereed) Published
Abstract [en]

High concentration of ascorbic acid (vitamin C) has been found in corneal epithelium of various species. However, the specific functions and mechanisms of ascorbic acid in the repair of corneal epithelium are not clear. In this study, it was found that ascorbic acid accelerates corneal epithelial wound healing in vivo in mouse. In addition, ascorbic acid enhanced the stemness of cultured mouse corneal epithelial stem/progenitor cells (TKE2) in vitro, as shown by elevated clone formation ability and increased expression of stemness markers (especially p63 and SOX2). The contribution of ascorbic acid on the stemness enhancement was not dependent on the promotion of Akt phosphorylation, as concluded by using Akt inhibitor, nor was the stemness found to be dependent on the regulation of oxidative stress, as seen by the use of two other antioxidants (GMEE and NAC). However, ascorbic acid was found to promote extracellular matrix (ECM) production, and by using two collagen synthesis inhibitors (AzC and CIS), the increased expression of p63 and SOX2 by ascorbic acid was decreased by around 50%, showing that the increased stemness by ascorbic acid can be attributed to its regulation of ECM components. Moreover, the expression of p63 and SOX2 was elevated when TKE2 cells were cultured on collagen I coated plates, a situation that mimics the in vivo situation as collagen I is the main component in the corneal stroma. This study shows direct therapeutic benefits of ascorbic acid on corneal epithelial wound healing and provides new insights into the mechanisms involved.

Place, publisher, year, edition, pages
WILEY, 2017
Keywords
Stem/progenitor cell, Colony formation, Proliferation, Microenvironment, Stem cell- croenvironment interactions, Tissue regeneration
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-136201 (URN)10.1002/sctm.16-0441 (DOI)000400596200009 ()28276172 (PubMedID)
Available from: 2017-07-03 Created: 2017-07-03 Last updated: 2019-02-22Bibliographically approved
Di, G., Qi, X., Zhao, X., Zhang, S., Danielson, P. & Zhou, Q. (2017). Corneal Epithelium-Derived Neurotrophic Factors Promote Nerve Regeneration. Investigative Ophthalmology and Visual Science, 58(11), 4695-4702
Open this publication in new window or tab >>Corneal Epithelium-Derived Neurotrophic Factors Promote Nerve Regeneration
Show others...
2017 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 58, no 11, p. 4695-4702Article in journal (Refereed) Published
Abstract [en]

PURPOSE. To explore the neurotrophic factor expression in corneal epithelium and evaluate their effects on the trigeminal ganglion (TG) neurite outgrowth and corneal nerve regeneration in mice. METHODS. The expression of neurotrophic factors was compared among the intact, regenerating, and regenerated mouse corneal epithelium. Mouse primary TG neurons were treated with the conditioned medium of mouse corneal epithelial cells. Nerve growth factor (NGF) neutralizing antibody and glial cell-derived neurotrophic factor (GDNF) neutralizing antibody were used to evaluate their roles in mouse corneal nerve regeneration and TG neurite outgrowth. The promoting effects of NGF and GDNF for the corneal nerve regeneration were further evaluated in the diabetic mice. RESULTS. The expression of NGF and GDNF showed significant up-regulation in regenerating corneal epithelium and return to the preinjury levels in the regenerated epithelium, which was consistent with the progress of corneal subbasal nerve regeneration. The conditioned medium of corneal epithelial cells promoted the TG neurite outgrowth with extended branching and elongation. Furthermore, the blockage of either NGF or GDNF significantly impaired the promotion of the neurite outgrowth by the conditioned medium or the corneal nerve regeneration in normal mice. Moreover, the expression of NGF and GDNF was attenuated in the diabetic regenerating corneal epithelium as compared to that in normal mice, while exogenous NGF or GDNF supplement promoted the corneal epithelial and nerve regeneration in diabetic mice. CONCLUSIONS. Corneal epithelium expresses multiple neurotrophic factors, among which NGF and GDNF may play an important role in the corneal nerve regeneration.

Place, publisher, year, edition, pages
ASSOC RESEARCH VISION OPHTHALMOLOGY INC, 2017
Keywords
corneal epithelium, neurotrophic factors, nerve regeneration, diabetes
National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-140476 (URN)10.1167/iovs.16-21372 (DOI)000410944300032 ()28910445 (PubMedID)
Available from: 2017-10-23 Created: 2017-10-23 Last updated: 2018-06-09Bibliographically approved
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
Show others...
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
Chen, J., Zhang, W., Kelk, P., Backman, L. J. & Danielson, P. (2017). Substance P and patterned silk biomaterial stimulate periodontal ligament stem cells to form corneal stroma in a bioengineered three-dimensional model. Stem Cell Research & Therapy, 8, Article ID 260.
Open this publication in new window or tab >>Substance P and patterned silk biomaterial stimulate periodontal ligament stem cells to form corneal stroma in a bioengineered three-dimensional model
Show others...
2017 (English)In: Stem Cell Research & Therapy, E-ISSN 1757-6512, Vol. 8, article id 260Article in journal (Refereed) Published
Abstract [en]

Background: We aimed to generate a bioengineered multi-lamellar human corneal stroma tissue in vitro by differentiating periodontal ligament stem cells (PDLSCs) towards keratocytes on an aligned silk membrane.

Methods: Human PDLSCs were isolated and identified. The neuropeptide substance P (SP) was added in keratocyte differentiation medium (KDM) to evaluate its effect on keratocyte differentiation of PDLSCs. PDLSCs were then seeded on patterned silk membrane and cultured with KDM and SP. Cell alignment was evaluated and the expression of extracellular matrix (ECM) components of corneal stroma was detected. Finally, multi-lamellar tissue was constructed in vitro by PDLSCs seeded on patterned silk membranes, which were stacked orthogonally and stimulated by KDM supplemented with SP for 18 days. Sections were prepared and subsequently stained with hematoxylin and eosin or antibodies for immunofluorescence observation of human corneal stroma-related proteins.

Results: SP promoted the expression of corneal stroma-related collagens (collagen types I, III, V, and VI) during the differentiation induced by KDM. Patterned silk membrane guided cell alignment of PDLSCs, and important ECM components of the corneal stroma were shown to be deposited by the cells. The constructed multi-lamellar tissue was found to support cells growing between every two layers and expressing the main type of collagens (collagen types I and V) and proteoglycans (lumican and keratocan) of normal human corneal stroma.

Conclusions: Multi-lamellar human corneal stroma-like tissue can be constructed successfully in vitro by PDLSCs seeded on orthogonally aligned, multi-layered silk membranes with SP supplementation, which shows potential for future corneal tissue engineering.

Place, publisher, year, edition, pages
BIOMED CENTRAL LTD, 2017
Keywords
PDLSCs, Corneal stroma, Substance P, Aligned silk membrane, Differentiation
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-142239 (URN)10.1186/s13287-017-0715-y (DOI)000415012400001 ()
Available from: 2017-12-11 Created: 2017-12-11 Last updated: 2019-02-22Bibliographically approved
Zhang, W., Chen, J., Backman, L. J., Malm, A. D. & Danielson, P. (2017). Surface Topography and Mechanical Strain Promote Keratocyte Phenotype and Extracellular Matrix Formation in a Biomimetic 3D Corneal Model. Advanced Healthcare Materials, 6(5), Article ID UNSP 1601238.
Open this publication in new window or tab >>Surface Topography and Mechanical Strain Promote Keratocyte Phenotype and Extracellular Matrix Formation in a Biomimetic 3D Corneal Model
Show others...
2017 (English)In: Advanced Healthcare Materials, ISSN 2192-2640, E-ISSN 2192-2659, Vol. 6, no 5, article id UNSP 1601238Article in journal (Refereed) Published
Abstract [en]

The optimal functionality of the native corneal stroma is mainly dependent on the well-ordered arrangement of extracellular matrix (ECM) and the pressurized structure. In order to develop an in vitro corneal model, it is crucial to mimic the in vivo microenvironment of the cornea. In this study, the influence of surface topography and mechanical strain on keratocyte phenotype and ECM formation within a biomimetic 3D corneal model is studied. By modifying the surface topography of materials, it is found that patterned silk fibroin film with 600 grooves mm(-1) optimally supports cell alignment and ECM arrangement. Furthermore, treatment with 3% dome-shaped mechanical strain, which resembles the shape and mechanics of native cornea, significantly enhances the expression of keratocyte markers as compared to flat-shaped strain. Accordingly, a biomimetic 3D corneal model, in the form of a collagen-modified, silk fibroin-patterned construct subjected to 3% dome-shaped strain, is created. Compared to traditional 2D cultures, it supports a significantly higher expression of keratocyte and ECM markers, and in conclusion better maintains keratocyte phenotype, alignment, and fusiform cell shape. Therefore, the novel biomimetic 3D corneal model developed in this study serves as a useful in vitro 3D culture model to improve current 2D cultures for corneal studies.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-134745 (URN)10.1002/adhm.201601238 (DOI)000399716300010 ()
Available from: 2017-05-11 Created: 2017-05-11 Last updated: 2019-02-22Bibliographically approved
Fong, G., Backman, L. J., Alfredson, H., Scott, A. & Danielson, P. (2017). The Effects of Substance P and Acetylcholine on Human Tenocyte Proliferation Converge Mechanistically via TGF-β1. PLoS ONE, 12(3), Article ID e0174101.
Open this publication in new window or tab >>The Effects of Substance P and Acetylcholine on Human Tenocyte Proliferation Converge Mechanistically via TGF-β1
Show others...
2017 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, no 3, article id e0174101Article in journal (Refereed) Published
Abstract [en]

Previous in vitro studies on human tendon cells (tenocytes) have demonstrated that the exogenous administration of substance P (SP) and acetylcholine (ACh) independently result in tenocyte proliferation, which is a prominent feature of tendinosis. Interestingly, the possible link between SP and ACh has not yet been explored in human tenocytes. Recent studies in other cell types demonstrate that both SP and ACh independently upregulate TGF-β1 expression via their respective receptors, the neurokinin 1 receptor (NK-1R) and muscarinic ACh receptors (mAChRs). Furthermore, TGF-β1 has been shown to downregulate NK-1R expression in human keratocytes. The aim of this study was to examine if TGF-β1 is the intermediary player involved in mediating the proliferative pathway shared by SP and ACh in human tenocytes. The results showed that exogenous administration of SP and ACh both caused significant upregulation of TGF-β1 at the mRNA and protein levels. Exposing cells to TGF-β1 resulted in increased cell viability of tenocytes, which was blocked in the presence of the TGFβRI/II kinase inhibitor. In addition, the proliferative effects of SP and ACh on tenocytes were reduced by the TGFβRI/II kinase inhibitor; this supports the hypothesis that the proliferative effects of these signal substances are mediated via the TGF-β axis. Furthermore, exogenous TGF-β1 downregulated NK-1R and mAChRs expression at both the mRNA and protein levels, and these effects were negated by simultaneous exposure to the TGFβRI/II kinase inhibitor, suggesting a negative feedback loop. In conclusion, the results indicate that TGF-β1 is the intermediary player through which the proliferative actions of both SP and ACh converge mechanistically.

National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-131396 (URN)10.1371/journal.pone.0174101 (DOI)000396318300126 ()
Note

Originally published in manuscript form

Available from: 2017-02-13 Created: 2017-02-13 Last updated: 2019-02-22Bibliographically approved
Organisations

Search in DiVA

Show all publications