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Sloniecka, Marta
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Publications (10 of 16) Show all publications
Sloniecka, M. & Danielson, P. (2019). Substance P induces fibrotic changes through activation of the RhoA/ROCK pathway in an in vitro human corneal fibrosis model. Journal of Molecular Medicine, 97(10), 1477-1489
Open this publication in new window or tab >>Substance P induces fibrotic changes through activation of the RhoA/ROCK pathway in an in vitro human corneal fibrosis model
2019 (English)In: Journal of Molecular Medicine, ISSN 0946-2716, E-ISSN 1432-1440, Vol. 97, no 10, p. 1477-1489Article in journal (Refereed) Published
Abstract [en]

Fibrosis is characterized by hardening, overgrowth, and development of scars in various tissues as a result of faulty reparative processes, diseases, or chronic inflammation. During the fibrotic process in the corneal stroma of the eye, the resident cells called keratocytes differentiate into myofibroblasts, specialized contractile fibroblastic cells that produce excessive amounts of disorganized extracellular matrix (ECM) and pro-fibrotic components such as alpha-smooth muscle actin (alpha-SMA) and fibronectin. This study aimed to elucidate the role of substance P (SP), a neuropeptide that has been shown to be involved in corneal wound healing, in ECM production and fibrotic markers expression in quiescent human keratocytes, and during the onset of fibrosis in corneal fibroblasts, in an in vitro human corneal fibrosis model. We report that SP induces keratocyte contraction and upregulates gene expression of collagens I, III, and V, and fibrotic markers: alpha-SMA and fibronectin, in keratocytes. Using our in vitro human corneal fibrosis model, we show that SP enhances gene expression and secretion of collagens I, III, and V, and lumican. Moreover, SP upregulates gene expression and secretion of alpha-SMA and fibronectin, and increases contractility of corneal fibroblasts during the onset of fibrosis. Activation of the preferred SP receptor, the neurokinin-1 receptor (NK-1R), is necessary for the SP-induced pro-fibrotic changes. In addition, SP induces the pro-fibrotic changes through activation of the RhoA/ROCK pathway. Taken together, we show that SP has a pro-fibrotic effect in both quiescent human keratocytes and during the onset of fibrosis in an in vitro human corneal fibrosis model.

Place, publisher, year, edition, pages
Springer Berlin/Heidelberg, 2019
Keywords
Cornea, Keratocytes, Scarring, Collagens, Fibrotic markers
National Category
Ophthalmology
Identifiers
urn:nbn:se:umu:diva-164380 (URN)10.1007/s00109-019-01827-4 (DOI)000486177200009 ()31399750 (PubMedID)
Available from: 2019-11-12 Created: 2019-11-12 Last updated: 2019-11-12Bibliographically approved
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
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
El-Habta, R., Sloniecka, M., Kingham, P. J. & Backman, L. J. (2018). The adipose tissue stromal vascular fraction secretome enhances the proliferation but inhibits the differentiation of myoblasts. Stem Cell Research & Therapy, 9, Article ID 352.
Open this publication in new window or tab >>The adipose tissue stromal vascular fraction secretome enhances the proliferation but inhibits the differentiation of myoblasts
2018 (English)In: Stem Cell Research & Therapy, E-ISSN 1757-6512, Vol. 9, article id 352Article in journal (Refereed) Published
Abstract [en]

Background: Adipose tissue is an excellent source for isolation of stem cells for treating various clinical conditions including injuries to the neuromuscular system. Many previous studies have focused on differentiating these adipose stem cells (ASCs) towards a Schwann cell-like phenotype (dASCs), which can enhance axon regeneration and reduce muscle atrophy. However, the stromal vascular fraction (SVF), from which the ASCs are derived, also exerts broad regenerative potential and might provide a faster route to clinical translation of the cell therapies for treatment of neuromuscular disorders.

Methods: The aim of this study was to establish the effects of SVF cells on the proliferation and differentiation of myoblasts using indirect co-culture experiments. A Growth Factor PCR Array was used to compare the secretomes of SVF and dASCs, and the downstream signaling pathways were investigated.

Results: SVF cells, unlike culture-expanded dASCs, expressed and secreted hepatocyte growth factor (HGF) at concentrations sufficient to enhance the proliferation of myoblasts. Pharmacological inhibitor studies revealed that the signal is mediated via ERK1/2 phosphorylation and that the effect is significantly reduced by the addition of 100 pM Norleual, a specific HGF inhibitor. When myoblasts were differentiated into multinucleated myotubes, the SVF cells reduced the expression levels of fast-type myosin heavy chain (MyHC2) suggesting an inhibition of the differentiation process.

Conclusions: In summary, this study shows the importance of HGF as a mediator of the SVF effects on myoblasts and provides further evidence for the importance of the secretome in cell therapy and regenerative medicine applications.

Place, publisher, year, edition, pages
BioMed Central, 2018
Keywords
Adipose stem cells, Differentiation, HGF, Myoblasts, Myotubes, Myogenesis, Proliferation, SVF
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-155097 (URN)10.1186/s13287-018-1096-6 (DOI)000454165400001 ()30572954 (PubMedID)
Available from: 2019-01-10 Created: 2019-01-10 Last updated: 2019-11-25Bibliographically 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
Sloniecka, M., Backman, L. J. & Danielson, P. (2016). Antiapoptotic Effect of Acetylcholine in Fas-Induced Apoptosis in Human Keratocytes. Investigative Ophthalmology and Visual Science, 57(14), 5892-5902
Open this publication in new window or tab >>Antiapoptotic Effect of Acetylcholine in Fas-Induced Apoptosis in Human Keratocytes
2016 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 57, no 14, p. 5892-5902Article in journal (Refereed) Published
Abstract [en]

PURPOSE. To investigate the possible antiapoptotic effect of acetylcholine (ACh) in Fas-mediated apoptosis of primary human keratocytes in vitro, and to explore the underlying mechanism. METHODS. Primary human keratocytes were isolated from healthy corneas. Fas ligand (FasL) was used to induce apoptosis in keratocytes. Cell death was assessed by ELISA. Activity of caspase-3, -7, -8, and -9 was measured with luminescent caspase activity assays. Expression of nuclear factor-kappa B (NF-kappa B) gene was assessed with RT-quantitative (q)PCR. Cytochrome c release apoptosis assay kit was used to extract mitochondria and cytosol. Cytochrome c release, cleavage of Bid, and expression of B-cell lymphoma 2 (Bcl-2) were determined by Western blot. RESULTS. Cell death ELISA revealed that ACh is able to reduce Fas-induced apoptosis in keratocytes. Analysis of the activity of effector caspases-3 and -7 showed that ACh, when added to Fas-treated cells, decreases the activation of both these enzymes. The activity of initiator caspases -8 and -9 also decreased when ACh was added to Fas-treated cells. This antiapoptotic effect of ACh was dependent on ACh concentration and activation of muscarinic ACh receptors. Analysis of the antiapoptotic mechanisms triggered by ACh showed that ACh downregulates expression of FasL-induced NF-kappa B RNA expression, upregulates expression of antiapoptotic protein Bcl-2, downregulates expression of proapoptotic protein Bad, reduces cytochrome c release, and prevents proapoptotic Bid protein cleavage. CONCLUSIONS. Acetylcholine has an antiapoptotic effect in a Fas-apoptosis model of human primary keratocytes in vitro. It is therefore possible that ACh may play a role in corneal wound healing, by modulating its initiation phase.

Keywords
Bcl-2, cytochrome c, muscarinic acetylcholine receptors, caspases, cornea
National Category
Cell and Molecular Biology Ophthalmology
Identifiers
urn:nbn:se:umu:diva-131665 (URN)10.1167/iovs.16-19707 (DOI)000392380000006 ()27802519 (PubMedID)
Available from: 2017-02-24 Created: 2017-02-24 Last updated: 2019-02-22Bibliographically approved
Erttmann, S. F., Härtlova, A., Sloniecka, M., Raffi, F. A. M., Hosseinzadeh, A., Edgren, T., . . . Gekara, N. O. (2016). Loss of the DNA Damage Repair Kinase ATM Impairs Inflammasome-Dependent Anti-Bacterial Innate Immunity. Immunity, 45(1), 106-118
Open this publication in new window or tab >>Loss of the DNA Damage Repair Kinase ATM Impairs Inflammasome-Dependent Anti-Bacterial Innate Immunity
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2016 (English)In: Immunity, ISSN 1074-7613, E-ISSN 1097-4180, Vol. 45, no 1, p. 106-118Article in journal (Refereed) Published
Abstract [en]

The ATM kinase is a central component of the DNA damage repair machinery and redox balance. ATM dysfunction results in the multisystem disease ataxia-telangiectasia (AT). A major cause of mortality in AT is respiratory bacterial infections. Whether ATM deficiency causes innate immune defects that might contribute to bacterial infections is not known. Here we have shown that loss of ATM impairs inflammasome- dependent anti-bacterial innate immunity. Cells from AT patients or Atm(-/-) mice exhibited diminished interleukin-1 beta (IL-1 beta) production in response to bacteria. In vivo, Atm(-/-) mice were more susceptible to pulmonary S. pneumoniae infection in a manner consistent with inflammasome defects. Our data indicate that such defects were due to oxidative inhibition of inflammasome complex assembly. This study reveals an unanticipated function of reactive oxygen species (ROS) in negative regulation of inflammasomes and proposes a theory for the notable susceptibility of AT patients to pulmonary bacterial infection.

National Category
Immunology in the medical area
Identifiers
urn:nbn:se:umu:diva-125590 (URN)10.1016/j.immuni.2016.06.018 (DOI)000380749000014 ()27421701 (PubMedID)
Available from: 2016-09-23 Created: 2016-09-13 Last updated: 2018-06-07Bibliographically 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
Słoniecka, M., Backman, L. J. & Danielson, P. (2015). Acetylcholine enhances keratocyte proliferation through muscarinic receptor activation.. International Immunopharmacology, 29(1), 57-62
Open this publication in new window or tab >>Acetylcholine enhances keratocyte proliferation through muscarinic receptor activation.
2015 (English)In: International Immunopharmacology, ISSN 1567-5769, E-ISSN 1878-1705, Vol. 29, no 1, p. 57-62Article in journal (Refereed) Published
Abstract [en]

Acetylcholine (ACh), a classical neurotransmitter, has been shown to be present in various non-neuronal cells, including cells of the eye, such as corneal epithelium and endothelium, and to have widespread physiological effects such as cytoskeleton reorganization, cellular proliferation, differentiation, and apoptosis. The aim of this study was to investigate the effect of ACh on corneal keratocyte proliferation, and the underlying mechanisms, in order to explore its possible effect in corneal wound healing. Primary culture of human keratocytes was established from donated corneas. Cell viability and fraction of proliferating cells were detected by MTS assay and BrdU incorporation ELISA, respectively. Expression of proliferative markers, PCNA and Ki-67, was detected by western blot and immunocytochemistry. Activation of the MAPK/Erk signaling pathway and its involvement in ACh-enhanced proliferation was determined by western blot analysis, MTS, and BrdU ELISA. We found that ACh enhanced keratocyte proliferation even at low concentrations. Stimulation of proliferation was mediated through activation of muscarinic ACh receptors (mAChRs). Western blot analysis revealed that ACh stimulation of keratocytes upregulated the expression of PCNA and Ki-67, and Ki-67 immunocytochemistry showed that ACh-treated cells were in an active phase of the cell cycle. ACh activated MAPK signaling, and this step was crucial for the ACh-enhanced proliferation, as inhibition of the MAPK pathway resulted in ACh having no proliferative effect. In conclusion, ACh enhances keratocyte proliferation and might thus play a role in proper corneal wound healing.

Keywords
substance P, acetylcholine, migration, proliferation, apoptosis, corneal stroma
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-112026 (URN)10.1016/j.intimp.2015.05.039 (DOI)000365365500010 ()26049030 (PubMedID)2-s2.0-84946489978 (Scopus ID)
Available from: 2015-11-30 Created: 2015-11-30 Last updated: 2019-02-22Bibliographically approved
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