Skin Fibroblast Cell Migration Research Articles

Electrospun hydroxypropyl-β-cyclodextrin nanofibers loaded with anti-inflammatory drugs for potential therapeutic wound dressings

Electrospun cyclodextrin nanofibers as innovative drug delivery structures offer efficient, selective, and controlled drug release. In the present work, free-standing nanofibrous webs (NFs) were fabricated from inclusion complexes (ICs) between 2-hydroxypropyl-β-cyclodextrin (HPβCD) and non-steroidal anti-inflammatory drugs (NSAIDs) using an electrospinning technique. The fabricated NSAID-HPβCD-IC-NFs were characterized by field emission scanning electron microscopy, X-ray diffraction, thermogravimetric/differential thermal analysis, ultraviolet diffuse reflectance spectroscopy, and photoluminescence analysis. Fourier transform infrared spectroscopy, 1H nuclear magnetic resonance spectroscopy, and computational studies were carried out to characterize the NSAID-HPβCD-IC-NFs and identify the functional groups involved in the binding mechanism. The NFs showed higher thermal stability than the pure drugs due to IC formation. The ultraviolet–visible absorption and fluorescence results indicated strong interactions between the HPβCD and NSAIDs with significant association constants at 1:1 stoichiometry. The efficiency of the NFs in terms of cell biocompatibility was verified, and the NFs exhibited excellent biocompatibility, as determined by in vitro biocompatibility assay. The in vitro scratch assay confirmed that the NFs allow the rapid growth and migration of skin fibroblast cells. These results showed that these NFs could be potential candidates for wound treatment.

Investigation of the functions of n-3 very-long-chain PUFAs in skin using in vivo Atlantic salmon and in vitro human and fish skin models.

The purpose of this study was to investigate the effect of dietary n-3 very-long-chain PUFA (n-3 VLC-PUFA) on the maturation and development of skin tissue in juvenile Atlantic salmon (Salmo salar) in vivo, as well as their effects on skin keratocyte and human skin fibroblast cell migration in vitro. Atlantic salmon were fed different dietary levels of n-3 VLC-PUFA from an initial weight of 6 g to a final weight of 11 g. Changes in skin morphology were analysed at two time points during the experiment, and the effects on skin tissue fatty acid composition were determined. Additionally, in vitro experiments using human dermal fibroblasts and primary Atlantic salmon keratocytes were conducted to investigate the effect of VLC-PUFA on the migration capacity of the cells. The results demonstrated that increased dietary levels of n-3 VLC-PUFA led to an increased epidermis thickness and more rapid scale maturation in Atlantic salmon skin in vivo, leading to a more mature skin morphology, and possibly more robust skin, at an earlier life stage. Additionally, human skin fibroblasts and salmon skin keratocytes supplemented with n-3 VLC-PUFA in vitro showed more rapid migration, indicating potentially beneficial effects of VLC-PUFA in wound healing. In conclusion, VLC-PUFA may have beneficial effects on skin tissue development, function and integrity.

Hyaluronic acid-quercetin pendant drug conjugate for wound healing applications

In this study, a simple approach was used for the synthesis of a water-soluble hyaluronic acid-quercetin (HA-Q) pendant drug conjugate to evaluate its potential wound-healing properties. The HA-Q conjugation was confirmed by Fourier-transform infrared spectroscopy (FTIR), ultraviolet–visible spectrophotometry (UV–Vis), and nuclear magnetic resonance (NMR) spectroscopy techniques. To produce the HA-Q, quercetin was conjugated on the HA backbone to the extent of 44.7 %. The HA-Q conjugate was soluble in water and a solution with a concentration of 20 mg/ml was prepared. The conjugate exhibited good biocompatibility and supported the growth and cell migration of skin fibroblast cells. HA-Q presented improved radical scavenging capacity compared to quercetin (Q) alone. The overall results confirmed the potential role of HA-Q in wound healing applications.

GSK3β-Ikaros-ANXA4 signaling inhibits high-glucose-induced fibroblast migration

Previous studies showed that the activation of Wnt signaling reduced high glucose (HG)-mediated fibroblast damage, but the molecular basis for this phenomenon remains elusive. This study aimed to analyze the level of phosphorylation of GSK3β Ser9 (pGSK3β Ser9) during HG damage. Moreover, the phosphomimic form of pGSK3β Ser9 was expressed to analyze its effect on cell migration via the phosphorylation of Ikaros. The results revealed that HG treatment significantly reduced the pGSK3β Ser9 level. The overexpression of GSK3β Ser9D and GSK3β Ser9A accelerated and inhibited fibroblast cell migration, respectively. P110α knockdown or treatment with SP600125, an inhibitor of JNK, also reduced the pGSK3β Ser9 level under HG condition. Treatment with SP600125 inhibited the migration of fibroblasts, but not in GSK3β Ser9D–expressing cells. Further, yeast two-hybrid screening and biochemical analysis identified that GSK3β interacted and phosphorylated Ikaros at Ser391. Besides, GSK3β Ser9D, but not GSK3β Ser9A, activated Ikaros Ser391 phosphorylation. Expressing Ikaros or β-catenin significantly promoted cell migration, suggesting that GSK3β modulated cell migration partially via the activation of Ikaros besides β-catenin signaling under HG condition. The expression of the phosphomimic form of Ikaros Ser391D resulted in a significant increase in the extent of cell migration compared with Ikaros under HG condition. Moreover, the Ikaros Ser391D DNA-binding affinity toward the ANXA4 promoter increased, and ANXA4 suppression promoted cell migration. In conclusion, the results of this study provided a new regulatory mechanism by which GSK3β negatively regulated human skin fibroblast cell migration.

Adipose stem cells isolated from diabetic mice improve cutaneous wound healing in streptozotocin-induced diabetic mice

BackgroundAdipose-derived mesenchymal stem cells (ASCs) therapy is emerging as a novel therapeutic option for the treatment of a variety of diseases including diabetes and diabetic wound healing. Multiple studies indicate that ASCs could promote wound healing and reverse diabetes. However, whether ASCs from diabetic donors retain their therapeutic functions and the mechanisms of how ASCs contribute to wound healing remain largely unknown. In this study, we explored the cutaneous wound healing ability of ASCs collected from C57BL/6 mice that had been rendered diabetic with streptozotocin (STZ).MethodsASCs were harvested from adipose tissues of type 1 diabetic (T1D) or normal C57BL/6 mice. Cell phenotypes were evaluated by flow cytometry analysis, and cell differentiation into adipocytes, chondrocytes, and osteocytes was compared. Secretions of transforming growth factor β (TGF-β1), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) by ASCs were assessed by ELISA. Migration and proliferation of fibroblasts co-cultured with T1D ASCs or control ASCs were also compared. The therapeutic effects of T1D and control ASCs in promoting wound closure were measured in vivo in a T1D wound mouse model. Granulation tissues were collected and stained with H&E at 14th day. CD34 and collagen I were detected by immunohistochemistry. Expressions of IL-6, α-SMA, CD31, collagen I, and collagen III were quantified by real-time PCR. GFP-expressing ASCs were used to trace in vivo cell differentiation.ResultsT1D ASCs and control ASCs showed similar expression of cell surface markers (CD29, CD34, CD105) and proliferation pattern. They can both differentiate into different cell types. T1D ASCs secreted similar amounts of VEGF and bFGF, but less TGF-β compared with control ASCs. Like control ASCs, T1D ASCs promoted the proliferation and migration of skin fibroblast cells. When injected in cutaneous wound of T1D mice, T1D ASCs increased wound closure and hair follicle regeneration at a comparable extent as ASCs. Mice receiving T1D ASCs or ASCs exhibited significantly higher expressions of collagen I, collagen III, and CD31 and reduced expression of IL-6 in wound tissues. Immunohistochemistry staining showed increased angiogenesis in mice receiving ASCs as was evident by increased CD34+ cells and collagen I staining. GFP+ ASCs injection showed that ASCs differentiated into fibroblasts and endothelial cells in vivo.ConclusionsOur results suggest that T1D ASCs could accelerate cutaneous wound healing. Mechanisms may include increasing fibroblast growth and migration, skin angiogenesis, and differentiation into fibroblasts and endothelial cells. This study provides evidence that diabetic ASCs may be used as a therapeutic option in cutaneous wound healing in diabetic recipients.

Bio-guided fractionation and isolation of active component from Tragopogon graminifolius based on its wound healing property

Ethnopharmacological relevanceTragopogon graminifolius (T. graminifolius) from Asteraceae family has been used as a remedy in Persian traditional medicine for the treatment of various disorders such as wound healing. Aim of the studyThe purpose of this study is to investigate the compounds of T. graminifolius, which are responsible for its wound healing activity. Materials and methodsThe experiment was performed in three phases; each phase consisted of fractionation of extracts followed by scratch assay. The results of the scratch assay were expressed using scratch closure index (SCI), representing the contraction of scratch. ResultsIn phase I, Ethyl acetate fraction (E) showed the maximum SCI (61.7 ± 3.5) that was selected for more fractionation in the next phase. In phase II, 12 fractions were obtained and labeled as fractions E- A to L, respectively. Based on the SCI of fractions, EF (SCI=68.9 ± 0.6) was the most active fraction in phase II and selected for further fractionation in phase III. In phase III, 8 fractions were resulted by fractionation of EF and labeled as EF- 1–8. Fraction EF5 with the highest SCI (30.8 ± 3.0) was the most effective fraction and Luteolin was the main component. Luteolin significantly improved viability of fibroblast cells and increased cell population that was accompanied by decreased cell apoptosis. Luteolin-induced cell number increase in the S and G2M phases of the cell cycle, further confirms the proliferative effect of this compound. ConclusionThe results showed that the total extract and fractions of T. graminifolius stimulate proliferation and migration of skin fibroblast cells and Luteolin is one of the active compounds responsible for these effects.

Annexin A2 functions downstream of c‑Jun N‑terminal kinase to promote skin fibroblast cell migration.

Delayed healing of skin wounds is one of the outcomes of diabetes mellitus (DM), a condition that affects a significant number of patients worldwide. However, the underlying mechanisms remain unknown. In order to examine proteome alterations in DM, a rat model of type 1 diabetes was developed using streptozotocin injections. The proteomic responses of normal and DM rat skin were analyzed by two‑dimensional electrophoresis, and differentially expressed proteins were identified using a liquid chromatography/mass spectrometry system. DM induced 36 and repressed 41 differentially expressed proteins, respectively. Altered proteins were involved in a number of biological processes, including RNA and protein metabolism, the tricarboxylic acid cycle, glycolysis, cytoskeleton regulation, hydrogen detoxification and calcium‑mediated signal transduction. In addition, overexpression of annexin A2, one of the signaling proteins altered by DM, accelerated the rate of human skin fibroblast cell migration. Application of SP600125, an inhibitor of a key regulator of cell migration c‑Jun N‑terminal kinase (JNK), inhibited the migration of normal cells. By contrast, SP600125 treatment did not inhibit the migration of annexin A2‑overexpressed cells, indicating that annexin A2 may function downstream of JNK. In conclusion, the results of the present study reveal the potential proteomic responses to DM in skin tissues, and demonstrate a positive functional role of annexin A2 in fibroblast cell migration.

Distribution of fibroblast growth factors and their roles in skin fibroblast cell migration

Fibroblast growth factor (FGF)2/basic FGF is a member of the fibroblast growth factor family. Its function in skin wound healing has been well-characterized. However, the function of other FGFs in skin tissues remains to be elucidated. In the present study, FGF expression patterns in heart, liver, skin and kidney tissues were analyzed. Notably, in contrast to other tissues, only four FGFs, FGF2, 7, 10 and 21, were dominant in the skin. To examine FGF function in the wound healing process, mouse NIH3T3 fibroblast cells were treated with FGF2, FGF10 and FGF21, and cell migration was monitored. The results revealed that FGF treatment promoted cell migration, which is an important step in wound healing. In addition, FGF treatment enhanced the activity of c-Jun N-terminal kinase (JNK), a key regulator of fibroblast cell migration. To analyze its role in cell migration, FGF7 was overexpressed in fibroblast cells via a lentivirus system; however, this did not change cell migration speed. FGF2, 7, 10 and 21 were highly expressed in skin tissue, and all except FGF7 regulated fibroblast cell migration and activated JNK. The results of the present study increase our understanding of the role of FGFs in skin wound healing.

Protective role of microRNA-29a in denatured dermis and skin fibroblast cells after thermal injury.

ABSTRACTOur previous study has suggested that downregulated microRNA (miR)-29a in denatured dermis might be involved in burn wound healing. However, the exact role of miR-29a in healing of burn injury still remains unclear. Here, we found that expression of miR-29a was notably upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury, and thereafter gradually downregulated compared with control group. By contrast, the expression of collagen, type I, alpha 2 (COL1A2) and vascular endothelial growth factor (VEGF-A) were first reduced and subsequently upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury. We further identified COL1A2 as a novel target of miR-29a, which is involved in type I collagen synthesis, and showed that miR-29a negatively regulated the expression level of COL1A2 in skin fibroblast cells. In addition, VEGF-A, another target gene of miR-29a, was also negatively mediated by miR-29a in skin fibroblast cells. Inhibition of miR-29a expression significantly promoted the proliferation and migration of skin fibroblast cells after thermal injury, and knockdown of COL1A2 and VEGF-A reversed the effects of miR-29a on the proliferation and migration of skin fibroblast cells. Furthermore, we found that Notch2/Jagged2 signaling was involved in miR-29a response to burn wound healing. Our findings suggest that downregulated miR-29a in denatured dermis may help burn wound healing in the later phase, probably via upregulation of COL1A2 and VEGF-A expression, which can further enhance type I collagen synthesis and angiogenesis.

Herbal formula Astragali Radix and Rehmanniae Radix exerted wound healing effect on human skin fibroblast cell line Hs27 via the activation of transformation growth factor (TGF-β) pathway and promoting extracellular matrix (ECM) deposition

Astragali Radix (AR) and Rehmanniae Radix (RR) have long been used in traditional Chinese Medicine and as the principal herbs in treating diabetic foot ulcer. In this study, we investigated the effect of NF3, which comprises of AR and RR in the ratio of 2:1(w/w), on skin fibroblast cell migration and the activation of selected genes and proteins related to wound healing.Human skin fibroblast cell line Hs27 was treated with NF3 at 4mg/ml for 24h, and in vitro scratch wound healing and quantitative cell migration assays were performed, respectively. The expression of transformation growth factor (TGF-β1) and bone morphogenetic protein 6 (BMP6) in Hs27 cells with or without NF3 treatment was analyzed by western blot analysis. In addition, the expression of a panel of genes involved in human TGF-β signaling pathway was analyzed in Hs27 cells upon NF3 treatment (4mg/ml, 24h) by quantitative real-time PCR (qRT-PCR). Furthermore, the expression of several genes and proteins associated with ECM synthesis was investigated by qRT-PCR analysis or/and ELISA techniques. The results suggested that NF3 promoted the migration of human skin fibroblast cells. Western blot analysis demonstrated that NF3 up-regulated TGF-β1 and BMP-6 synthesis. qRT-PCR analysis revealed that the expression of 26 genes in Hs27 cells was changed upon NF3 induction, including TGF-β superfamily ligands and down stream effectors genes, and genes involved in TGF/Smad pathway, and Ras/MAPK (non-Smad) pathway. Among the extracellular matrix (ECM)-related molecules, it was found that NF3 up-regulated the expression of type I and III collagens, fibronectin as well as TIMP-1, and down-regulated the MMP-9 expression in skin fibroblast cells. This study demonstrated that herb formula NF3 could enhance skin fibroblast cell migration and activated genes involved in TGF-β1 pathway. NF3 could regulate gene transcription for extracellular matrix synthesis via the Smad pathway, and gene transcription for cell motility via the Ras/MAPK (non-Smad) pathway.

EGFR-mediated expression of aquaporin-3 is involved in human skin fibroblast migration

AQP3 (aquaporin-3), known as an integral membrane channel in epidermal keratinocytes, facilitates water and glycerol movement into and out of the skin. Here, we demonstrate that AQP3 is also expressed in cultured human skin fibroblasts, which under normal wound healing processes migrate from surrounding tissues to close the wound. EGF (epidermal growth factor), which induced fibroblast migration, also induced AQP3 expression in a time- and dose-dependent manner. CuSO4 and NiCl2, previously known as AQP3 water transport inhibitors, as well as two other bivalent heavy metals Mn2+ and Co2+, inhibited EGF-induced cell migration in human skin fibroblasts. AQP3 knockdown by small interfering RNA inhibited EGF-induced AQP3 expression and cell migration. Furthermore, an EGFR (EGF receptor) kinase inhibitor, PD153035, blocked EGF-induced AQP3 expression and cell migration. MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase]/ERK inhibitor U0126 and PI3K (phosphoinositide 3-kinase) inhibitor LY294002 also inhibited EGF-induced AQP3 expression and cell migration. Collectively, our findings show for the first time that AQP3 is expressed in human skin fibroblasts and that EGF induces AQP3 expression via EGFR, PI3K and ERK signal transduction pathways. We have provided evidence for a novel role of AQP3 in human skin fibroblast cell migration, which occurs during normal wound healing.