Ceramide kinase is required for a normal eicosanoid response and the subsequent orderly migration of fibroblasts

Abstract

In these studies, the role of ceramide-1-phosphate (C1P) in the wound-healing process was investigated. Specifically, fibroblasts isolated from mice with the known anabolic enzyme for C1P, ceramide kinase (CERK), ablated (CERK−/− mice) and their wild-type littermates (CERK+/+) were subjected to in vitro wound-healing assays. Simulation of mechanical trauma of a wound by scratching a monolayer of fibroblasts from CERK+/+ mice demonstrated steadily increasing levels of arachidonic acid in a time-dependent manner in stark contrast to CERK−/− fibroblasts. This observed difference was reflected in scratch-induced eicosanoid levels. Similar, but somewhat less intense, changes were observed in a more complex system utilizing skin biopsies obtained from CERK-null mice. Importantly, C1P levels increased during the early stages of human wound healing correlating with the transition from the inflammatory stage to the peak of the fibroplasia stage (e.g., proliferation and migration of fibroblasts). Finally, the loss of proper eicosanoid response translated into an abnormal migration pattern for the fibroblasts isolated from CERK−/−. As the proper migration of fibroblasts is one of the necessary steps of wound healing, these studies demonstrate a novel requirement for the CERK-derived C1P in the proper healing response of wounds. In these studies, the role of ceramide-1-phosphate (C1P) in the wound-healing process was investigated. Specifically, fibroblasts isolated from mice with the known anabolic enzyme for C1P, ceramide kinase (CERK), ablated (CERK−/− mice) and their wild-type littermates (CERK+/+) were subjected to in vitro wound-healing assays. Simulation of mechanical trauma of a wound by scratching a monolayer of fibroblasts from CERK+/+ mice demonstrated steadily increasing levels of arachidonic acid in a time-dependent manner in stark contrast to CERK−/− fibroblasts. This observed difference was reflected in scratch-induced eicosanoid levels. Similar, but somewhat less intense, changes were observed in a more complex system utilizing skin biopsies obtained from CERK-null mice. Importantly, C1P levels increased during the early stages of human wound healing correlating with the transition from the inflammatory stage to the peak of the fibroplasia stage (e.g., proliferation and migration of fibroblasts). Finally, the loss of proper eicosanoid response translated into an abnormal migration pattern for the fibroblasts isolated from CERK−/−. As the proper migration of fibroblasts is one of the necessary steps of wound healing, these studies demonstrate a novel requirement for the CERK-derived C1P in the proper healing response of wounds. Healing of cutaneous wounds proceeds through four distinct, yet overlapping stages. These stages include hemostasis, inflammation, proliferation, and remodeling (1Goldberg S.R. Diegelmann R.F. Wound healing primer.Surg. Clin. North Am. 2010; 90: 1133-1146Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar). During hemostasis, blood vessels constrict to minimize bleeding. The inflammatory stage of wound healing is then initiated and consists of vasodilation, increased vascular permeability, and neutrophil and monocyte (macro­phage) infiltration to the wound site. This stage clears bacteria and cellular debris. The proliferation phase begins toward the end of the inflammatory stage and involves the migration and proliferation of fibroblasts and keratinocytes. This stage also includes the formation of new blood vessels, induced by migrating endothelial cells, resulting in the formation of abundant granulation tissue. The final stage of wound healing is the maturation and remodeling phase. In this stage, new epithelium is formed, and the collagen content in the wound is increased and deposited in an orderly fashion. The high level of collagen cross-linking in this stage leads to a healed wound with ∼80%; the strength of the original skin. Fibroblasts are one of the most important groups of cells taking part in the wound-healing process. During the proliferation stage of wound healing, the fibroblasts in the surrounding tissue are stimulated to proliferate followed by migration into the wound. Fibroblasts provide the necessary extracellular matrix promoting the migration of additional cell types necessary for the completion of the wound-healing process. Thereafter, the fibroblasts alter their phenotype into myofibroblasts. Contraction of the myofibroblasts induces the wound edges to conjoin, minimizing the exposed surface area (1Goldberg S.R. Diegelmann R.F. Wound healing primer.Surg. Clin. North Am. 2010; 90: 1133-1146Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar, 2Berry D.P. Harding K.G. Stanton M.R. Jasani B. Ehrlich H.P. Human wound contraction: collagen organization, fibroblasts, and myofibroblasts.Plast. Reconstr. Surg. 1998; 102: 124-131Crossref PubMed Scopus (148) Google Scholar). While not as predominant as in rodents, wound contraction still accounts for up to 88%; of the mechanism toward wound closure in humans (1Goldberg S.R. Diegelmann R.F. Wound healing primer.Surg. Clin. North Am. 2010; 90: 1133-1146Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar) indicating the relevance of these cells in human wound healing. Inhibition or loss of fibroblast migration leads to these cells remaining at the margins and thus causes abnormal wound healing (3Xu F. Zhang C. Graves D.T. Abnormal cell responses and role of TNF-α in impaired diabetic wound healing..Biomed. Res. Int. 2013; 754802PubMed Google Scholar). On the other hand, if there is increased, disorganized, or random migration of fibroblasts, the deposition of collagen would be uneven and unregulated, leading to hypertrophic scar formation or healed wounds with compromised strength (4Suganuma H. Sato A. Tamura R. Chida K. Enhanced migration of fibroblasts derived from lungs with fibrotic lesions.Thorax. 1995; 50: 984-989Crossref PubMed Scopus (80) Google Scholar, 5White E.S. Atrasz R.G. Dickie E.G. Aronoff D.M. Stambolic V. Mak T.W. Moore B.B. Peters-Golden M. Prostaglandin E2 inhibits fibroblast migration by E-prostanoid 2 receptor–mediated increase in PTEN activity.Am. J. Respir. Cell Mol. Biol. 2005; 32: 135-141Crossref PubMed Scopus (122) Google Scholar, 6Huang S.K. Peters-Golden M. Eicosanoid lipid mediators in fibrotic lung diseases: ready for prime time?.Chest. 2008; 133: 1442-1450Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar, 7Rieger G.M. Hein R. Adelmann-Grill B.C. Ruzicka T. Krieg T. Influence of eicosanoids on fibroblast chemotaxis and protein synthesis in vitro.J. Dermatol. Sci. 1990; 1: 347-354Abstract Full Text PDF PubMed Scopus (18) Google Scholar). Several published studies demonstrate the importance of eicosanoids, a class of inflammatory lipids, in the migration of fibroblasts (7Rieger G.M. Hein R. Adelmann-Grill B.C. Ruzicka T. Krieg T. Influence of eicosanoids on fibroblast chemotaxis and protein synthesis in vitro.J. Dermatol. Sci. 1990; 1: 347-354Abstract Full Text PDF PubMed Scopus (18) Google Scholar, 8Green J.A. Stockton R.A. Johnson C. Jacobson B.S. 5-Lipoxygenase and cyclooxygenase regulate wound closure in NIH/3T3 fibroblast monolayers.Am. J. Physiol. Cell Physiol. 2004; 287: C373-C383Crossref PubMed Scopus (22) Google Scholar, 9Kohyama T. Liu X. Kim H.J. Kobayashi T. Ertl R.F. Wen F-Q. Takizawa H. Rennard S.I. Prostacyclin analogs inhibit fibroblast migration.Am. J. Physiol. Lung Cell. Mol. Physiol. 2002; 283: L428-L432Crossref PubMed Scopus (34) Google Scholar, 10Nieves D. Moreno J.J. Hydroxyeicosatetraenoic acids released through the cytochrome P-450 pathway regulate 3T6 fibroblast growth.J. Lipid Res. 2006; 47: 2681-2689Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar, 11Su W-H. Cheng M-H. Lee W-L. Tsou T-S. Chang W-H. Chen C-S. Wang P-H. Nonsteroidal anti-inflammatory drugs for wounds: pain relief or excessive scar formation?.Mediators Inflamm. 2010; 2010: 413238Crossref PubMed Scopus (70) Google Scholar). Green et al. (8Green J.A. Stockton R.A. Johnson C. Jacobson B.S. 5-Lipoxygenase and cyclooxygenase regulate wound closure in NIH/3T3 fibroblast monolayers.Am. J. Physiol. Cell Physiol. 2004; 287: C373-C383Crossref PubMed Scopus (22) Google Scholar) demonstrated that products of 5-lipoxygenase and cyclooxygenase (COX) are important in the regulation of wound closure in NIH/3T3 cells, while Kohyama et al. (9Kohyama T. Liu X. Kim H.J. Kobayashi T. Ertl R.F. Wen F-Q. Takizawa H. Rennard S.I. Prostacyclin analogs inhibit fibroblast migration.Am. J. Physiol. Lung Cell. Mol. Physiol. 2002; 283: L428-L432Crossref PubMed Scopus (34) Google Scholar) demonstrated that prostacyclin analogs inhibit the migration of fibroblasts. Rieger et al. (7Rieger G.M. Hein R. Adelmann-Grill B.C. Ruzicka T. Krieg T. Influence of eicosanoids on fibroblast chemotaxis and protein synthesis in vitro.J. Dermatol. Sci. 1990; 1: 347-354Abstract Full Text PDF PubMed Scopus (18) Google Scholar) demonstrated that 5- and 12-hydroxyeicosatetranoic acid (HETE) cause a dose-dependent increase in the chemotaxis of fibroblasts. Su et al. (11Su W-H. Cheng M-H. Lee W-L. Tsou T-S. Chang W-H. Chen C-S. Wang P-H. Nonsteroidal anti-inflammatory drugs for wounds: pain relief or excessive scar formation?.Mediators Inflamm. 2010; 2010: 413238Crossref PubMed Scopus (70) Google Scholar) demonstrated that nonsteroidal anti-inflammatory drugs result in excessive scar formation due to enhanced fibroblast migration and proliferation. These studies demonstrate an important role for eicosanoids in the migration and proliferation of fibroblasts. Group IVA cytosolic phospholipase A2 (cPLA2α) is one of the major phospholipases activated in response to mechanical insults, which leads to the production of arachidonic acid (AA). AA is metabolized enzymatically to various eicosanoids (prostaglandins, leukotrienes, thromboxanes, etc.). Ceramide-1-phosphate (C1P) is a potent and of and published demonstrate the requirement for ceramide kinase C1P in the of AA in response to inflammatory A. M. J. et is a of cytosolic phospholipase Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, A. kinase and arachidonic acid Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, A. as a of IVA cytosolic phospholipase A2 and the of the enzyme with Biol. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar, M. IVA cytosolic phospholipase A2 and Lipid Res. Full Text Full Text PDF PubMed Scopus Google Scholar, is for the of IVA cytosolic phospholipase A2 and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). the that CERK-derived C1P the migration of fibroblasts due to role in eicosanoid demonstrate that loss of the of fibroblasts to eicosanoids in response to mechanical as as in wound This compromised eicosanoid in the migration of fibroblasts in a random The requirement of C1P for orderly fibroblast migration with the observed peak production of C1P in the fibroplasia stage of human wound healing. these demonstrate that the of CERK-derived C1P is for the proper eicosanoid response and migration of fibroblasts into a wound site. and and were obtained from The was a and the was a from to were by a and acid were from fibroblasts were isolated from or wild-type (CERK+/+) or in as is for the of IVA cytosolic phospholipase A2 and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The were in high with and at These cells were at the stage or for to or obtained from CERK-null mice and their wild-type littermates were on tissue in with and The cells were to the was from to and a was on the monolayer the of the with a The migration of the fibroblasts into the area was or obtained from CERK-null mice and their wild-type littermates were on tissue in with and and with AA as A. A. for of by of the system to Lipid Res. 50: Full Text Full Text PDF PubMed Scopus Google Scholar). The monolayer obtained the was for 2 in and The monolayer was then subjected to a with a was at the and AA was cells were a the were on and the was to and in the and of eicosanoids as cells were with and by in of followed by to a were from the remaining cells a and and as by et al. M. of high for the of ceramide-1-phosphate Lipid Res. 2010; Full Text Full Text PDF PubMed Scopus Google Scholar). to of the cells in of was The were with of C1P, and as the The was to the cell and for at the were to a new and in by and at for The in and at is a new to published M. of high for the of ceramide-1-phosphate Lipid Res. 2010; Full Text Full Text PDF PubMed Scopus Google Scholar) and was into the to proper of the The lipid thus obtained levels of as by the not and was in the of the C1P, and The were a on a system and a in and at for at a of at an was to C1P, and were from as X. X. S.K. et by a ceramide-1-phosphate protein 2013; PubMed Scopus Google Scholar, J.A. R. of eicosanoid synthesis in a of ceramide Lipid Res. 2013; Full Text Full Text PDF PubMed Scopus Google Scholar). to of and acid were The were with the eicosanoids E2 and were with and then The were to the Thereafter, the were with which was then to the Finally, the eicosanoids were with The was and the were in for The lipid thus obtained levels of as by the not and the eicosanoids were utilizing a was to the eicosanoids at a of at The was with acid for and then of was The was for the of was increased in a to to to to to and to The was to in a and then The eicosanoids were then a in were which be in The were as 1: and cell were on with in a and then for with with and were with and for in was by with at in and was with for in were then on with and to at in the Cell was by cells a were into cell and to of migration was by scratching cells with a was on a with a in a with were for at was to wound closure at and to cell Cell and for by the of the cell from at of and by the and was to obtained from CERK-null mice and their wild-type littermates were on tissue in with and The cells were to The monolayer obtained the was for 2 in and The monolayer was then subjected to a with a and the was with or the eicosanoids and The migration of the fibroblasts into the area was at and the closure of the area was with the of and and the of was and were into the of the of was by a of the tissue in a a the skin was and the was through the skin The was then and the and of the were to the skin a was from the and the was in and for in the lipid The and the were with and a On and one was and in Lipid was for the CERK+/+ and CERK−/− were obtained from as a from cells derived from these mice were for as by and C. B. A. R. D. M. C. et with impaired response to in ceramide 2008; PubMed Scopus Google Scholar). were at The and was by the in with the of the on the and of and the for the and of in and is in with of the and other and to is the as a with the and The of the for in with the human studies were the of the of of and was obtained from are as was groups other for of four or more groups was a with was In these a was to of the with were were In to an on the C1P the levels of C1P in in the and of mechanical As levels of the major of C1P, as as that of were observed in the CERK−/− cells, the importance of in the synthesis of were also observed for C1P, this is on and C1P were to be in the CERK−/− the wild-type the CERK-null fibroblasts demonstrated additional changes to the C1P content of mechanical trauma These demonstrate that a of the and C1P is but the in the levels of C1P is demonstrated that C1P by was a necessary in the and of AA is for the of IVA cytosolic phospholipase A2 and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). CERK-derived C1P a role in in response to mechanical trauma known to In to this the AA of in response to scratch-induced mechanical a wild-type demonstrated increasing AA in contrast to CERK−/− of mechanical trauma by scratching the the wild-type demonstrated a increase of while CERK−/− demonstrated a and increase in AA The that the and mechanical AA by fibroblasts was due to a in the of In to this the levels of wild-type and the CERK-null fibroblasts. In with J.A. R. of eicosanoid synthesis in a of ceramide Lipid Res. 2013; Full Text Full Text PDF PubMed Scopus Google there was difference in the level of the not the loss of the AA in response to mechanical trauma the of AA is the of eicosanoids, and the production of AA is the of eicosanoid In to the observed of the of AA in response to mechanical trauma translated to the level of eicosanoids, a with for the of AA of CERK−/− demonstrated a in and induced AA with CERK+/+ the demonstrated that the in AA in CERK−/− translated to synthesis of the and and the prostacyclin the was also observed to the products and the that and mechanical of AA which into eicosanoid CERK-derived C1P an important and necessary role in the of eicosanoids in response to the of fibroblasts. In to on eicosanoid production to a more complex the synthesis of these by the skin of CERK−/− and CERK+/+ mice in response to mechanical biopsies were obtained from the of wild-type and CERK-null mice and were to The eicosanoids into the were and was observed in and in the CERK-null mice with the skin of their wild-type littermates at In on and were observed the groups the synthesis of eicosanoids in response to of the skin of CERK-null mice was with that of the wild-type a predominant role for CERK-derived C1P in the wound response of the skin. studies demonstrate the importance of eicosanoids in fibroblast migration and proliferation an important of the proliferation stage and the remodeling of wound healing T. T. V. The wound healing an of the cellular and Int. Res. PubMed Scopus Google Scholar). that and skin derived from CERK−/− mice impaired eicosanoid response to the of migration as the CERK−/− demonstrated an activated fibroblast phenotype with CERK+/+ as by the of more by In to the role of CERK-derived C1P in a monolayer of was subjected to scratch-induced mechanical trauma followed by of the cell that the CERK-null fibroblasts in a random pattern in contrast to their wild-type as by the CERK−/− a loss in migration the CERK-null fibroblasts demonstrated an increase in the wound area with wild-type fibroblasts these demonstrate that CERK-derived C1P a role in the proper migration of fibroblasts to a wound site. studies demonstrated that CERK-derived C1P is an important in the migration of fibroblasts induced by In to CERK-derived C1P fibroblast migration eicosanoid Specifically, the eicosanoids in the CERK−/− cells and skin were into the to the for wild-type fibroblasts. The demonstrate that while the not on the of migration of the CERK-null fibroblasts, of AA and the eicosanoids, and induced a in the migration of CERK−/− at the early of migration migration this wild-type and CERK-null fibroblasts subjected to surface were by in the and and These studies demonstrated a of the of wound closure by these in CERK-null fibroblasts at early of migration during this early the and the of the were the enhanced migration and of CERK-null fibroblasts is due to the production of In to the importance of C1P in fibroblast migration to wound healing in changes in the levels of this lipid in a human wound The levels of C1P were observed to increase from through through to These a increase in C1P through the inflammatory phase a on the peak of fibroplasia during the phase. there is a the levels of C1P and the migration stage of an important role for this lipid in fibroplasia during wound healing. In this a role for CERK-derived C1P in the wound-healing response of fibroblasts. Specifically, that of CERK-derived C1P to a AA and eicosanoid of fibroblasts in response to scratch-induced mechanical also demonstrated that this in changes to the and the pattern of migration of fibroblasts. for the a the CERK-derived C1P and process of fibroblasts. also the for a major role by CERK-derived C1P in the of important eicosanoid CERK-null mice were as C. M. B. F. levels of ceramide and ceramide-1-phosphate in the of ceramide Res. 2008; PubMed Scopus Google Scholar). of was observed to not the of in cell types as C. B. A. R. D. M. C. et with impaired response to in ceramide 2008; PubMed Scopus Google of these was that is not to eicosanoid in the of this CERK-derived of C1P and a is observed the of the levels of and C1P through an of synthesis as J.A. R. of eicosanoid synthesis in a of ceramide Lipid Res. 2013; Full Text Full Text PDF PubMed Scopus Google Scholar). for of C1P include an a enzyme with that from to C1P, or an enzyme with that C1P of this is not to the of the loss of on eicosanoid in Specifically, fibroblasts a of their to to as mechanical CERK-derived C1P to a role in this eicosanoid and this role be in of the importance of CERK-derived C1P in the AA response be for the cell and the and not be on the phenotype of a of cell types as impaired of fibroblasts to a eicosanoid response also for the other cells in the wound-healing process. demonstrated an fibroblasts and during wound healing Krieg T. H. in wound Dermatol. Full Text Full Text PDF PubMed Scopus Google Scholar). is known to proliferation of proliferation in vitro by Clin. PubMed Scopus Google Scholar). fibroblasts with are as a to provide the eicosanoids, indicating the relevance for the M. fibroblasts as for human the to Dermatol. Full Text PDF PubMed Scopus (28) Google Scholar). in the eicosanoid of the fibroblasts most the proliferation, the of the wound-healing process. is the is also necessary for the of wound by and loss to wounds. during the of this a was published by Kim et al. C. A. K. M. J. M. Ceramide-1-phosphate migration of cells and endothelial for tissue 2013; PubMed Scopus (70) Google Scholar) a C1P and the migration of cells and endothelial cells with for tissue are in with their in to the C1P requirement for fibroblast but in that the of C1P in fibroblasts is to eicosanoids important for migration and cell in an and due to the in the migration of fibroblasts are not to wound healing. are other that are the result of excessive and of fibroblasts and collagen These include and as as and and regulate in various Clin. PubMed Scopus Google Scholar). of the fibroblasts isolated from demonstrate an increased (4Suganuma H. Sato A. Tamura R. Chida K. Enhanced migration of fibroblasts derived from lungs with fibrotic lesions.Thorax. 1995; 50: 984-989Crossref PubMed Scopus (80) Google proliferation, and increased collagen synthesis that is with a to S.K. Peters-Golden M. Eicosanoid lipid mediators in fibrotic lung diseases: ready for prime time?.Chest. 2008; 133: 1442-1450Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar). also observed a in C1P toward the end of the stage of wound healing in which would with a in and of collagen synthesis by fibroblasts. the and observed in the CERK-null fibroblasts is to the phenotype of fibroblasts isolated from the fibrotic lung S.K. Peters-Golden M. Eicosanoid lipid mediators in fibrotic lung diseases: ready for prime time?.Chest. 2008; 133: 1442-1450Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar). The in observed in CERK−/− cells is of importance as the phenotype of the fibroblasts obtained isolated from a role for in (e.g., excessive scar and formation this also that the migration of fibroblasts is but orderly high while in vitro wound-healing demonstrated closure of the wound enhanced but random migration of CERK-null fibroblasts, in the of loss of be a uneven migration of these cells also be to result in uneven collagen deposition with toward the strength of the healed wound. cellular C1P content be an important in the phenotype as as strength of healed and studies are to this new While the of the migration of with AA and is with published and a by was This eicosanoid is the of prostacyclin and was to be As was in as a by a role for in the migration of is known the role of eicosanoids in fibroblast migration and wound healing, these that studies be to role this eicosanoid in this process. Abnormal migration be due to in one or more eicosanoid by et al. E.S. Atrasz R.G. Dickie E.G. Aronoff D.M. Stambolic V. Mak T.W. Moore B.B. Peters-Golden M. Prostaglandin E2 inhibits fibroblast migration by E-prostanoid 2 receptor–mediated increase in PTEN activity.Am. J. Respir. Cell Mol. Biol. 2005; 32: 135-141Crossref PubMed Scopus (122) Google Scholar) demonstrated that lung fibroblast migration is by an E-prostanoid 2 mechanism that increased production and and of PTEN the of the of and the loss of PTEN was as a cause of enhanced migration of fibroblasts J. R. R. D. H. H. of the cell by of and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The of the migration phenotype of CERK-null fibroblasts by of be to the of this AA is the of the observed of the enhanced migration phenotype by AA is due to the of AA into this mechanism not the by is a that this of prostacyclin is to a more which is for the observed by et al. Lee R.F. of prostacyclin by in human of a potent of and enzyme Biol. Full Text PDF PubMed Google Scholar) demonstrated that be into with to The for also leads to the of of (9Kohyama T. Liu X. Kim H.J. Kobayashi T. Ertl R.F. Wen F-Q. Takizawa H. Rennard S.I. Prostacyclin analogs inhibit fibroblast migration.Am. J. Physiol. Lung Cell. Mol. Physiol. 2002; 283: L428-L432Crossref PubMed Scopus (34) Google Scholar) and as be to inhibit fibroblast migration as by Kohyama et al. (9Kohyama T. Liu X. Kim H.J. Kobayashi T. Ertl R.F. Wen F-Q. Takizawa H. Rennard S.I. Prostacyclin analogs inhibit fibroblast migration.Am. J. Physiol. Lung Cell. Mol. Physiol. 2002; 283: L428-L432Crossref PubMed Scopus (34) Google Scholar) through the the observed in the migration of CERK-null fibroblasts that is by the of eicosanoids is to the of In demonstrated that the loss of in a of C1P which in in of the eicosanoids by fibroblasts. In this in eicosanoids in the enhanced but random migration of fibroblasts of cell studies in complex wound-healing in humans and mice this role for CERK-derived C1P in this this the of and thus C1P, in the of wound healing as as The of for of the CERK−/− with wild-type and the Cell for with arachidonic acid ceramide-1-phosphate ceramide kinase cyclooxygenase cytosolic phospholipase A2 acid fibroblast E2 and