Effect Of Epidermal Growth Factor Research Articles

AnnexinA2 regulates glioma cell proliferation through the STAT3‑cyclinD1 pathway.

Annexin A2 (ANXA2) acts as a calcium‑dependent phospholipid‑binding protein that is widely expressed in vertebrate cells and has abnormally high expression in various tumor cells. However, the detailed molecular mechanism underlying the effects of ANXA2 on glioma cells remains unclear. The present study aimed to investigate the role and underlying molecular mechanisms of ANXA2 in glioma cell proliferation. The results revealed that knockdown of ANXA2 inhibited the proliferation of U251 and U87 glioma cell lines and decreased phosphorylated (p) signal transducer and activator of transcription 3 (STAT3)(Y705) and cyclin D1 expression, leading to impedance of the G1‑to‑S phase transition. Furthermore, it was suggested that ANXA2 may regulate pSTAT3(Y705) levels through direct binding with STAT3, thereby affecting STAT3‑cyclin D1 pathway‑mediated cell proliferation. When ANXA2 was re‑expressed in ANXA2‑knockdown cells, the expression of pSTAT3(Y705) and cyclin D1 was restored. Furthermore, overexpression of ANXA2 significantly promoted the proliferation of U251 cells, as determined by an MTT assay and a tumor formation assay in nude mice, but had no statistically significant effect on colony formation rate, cell cycle progression or the STAT3‑cyclin D1 pathway, suggesting that endogenous ANXA2 may be redundant. Additionally, the present study provided evidence that the overexpression of ANXA2 enhanced the expression of pSTAT3(Y705) in the presence of epidermal growth factor (EGF), indicating that the proliferation‑promoting effect of ANXA2 may be due to the accumulation and synergistic effect of paracrine EGF. Taken together, the present results indicated that ANXA2 may affect the proliferation of human glioma cells through the STAT3‑cyclin D1 pathway via direct interaction with STAT3 in U251 and U87 glioma cells. ANXA2 was redundant in this pathway, but positive synergy was revealed to exist between ANXA2 and EGF.

Maternal EGF limits bacterial translocation in the offspring: a model of enterally acquired Late Onset Sepsis

Abstract Late-onset neonatal sepsis (LOS), an important cause of morbidity and mortality in prematurely born infants, results from a bloodstream infection by bacteria of gut origin. It remains unclear how such bacteria translocate from the intestine, though it has been repeatedly observed that breastfed infants, particularly those fed mother’s own milk (MOM), have reduced risk of LOS compared to formula fed infants. Epidermal growth factor (EGF) promotes intestinal barrier function in infants and is present at high concentrations in breast milk post-partum and decreases throughout lactation. We found reduced concentrations of EGF in the stool of premature formula-fed infants compared to MOM-fed infants, and observed a similar decrease in EGF concentrations of stool of neonatal mice asynchronously cross-fostered (ACF) to dams that had delivered two weeks prior. LOS bloodstream isolates of E. coli colonized the tracts of all pups but translocated and disseminated systemically only in ACF mice resulting in bacteremia and rapid death. Oral gavage of recombinant EGF reduced bacteria translocation and prevented the development of systemic disease in ACF mice. Thus, disruption of maternally delivered EGF in ACF mice results in translocation of pathogens from the gut, and a sepsis-like disease. In conclusion we have identified a mechanism whereby gut-residing pathogens gain systemic access and have developed a novel animal model replicating this mechanism to explore the protective effect of breastmilk and EGF in LOS.

Epidermal growth factor protects against myocardial ischaemia reperfusion injury through activating Nrf2 signalling pathway

Alleviating the oxidant stress associated with myocardial ischaemia reperfusion has been demonstrated as a potential therapeutic approach to limit ischaemia reperfusion (I/R)-induced cardiac damage. It is reported that EGFR/erbB2 signalling is an important cardiac survival pathway in cardiac function and activation of EGFR has a cardiovascular effect in global ischaemia. Epidermal growth factor (EGF), a typical EGFR ligand, was considered to have a significant role in activating EGFR. However, no evidence has been published whether exogenous EGF has protective effects on myocardial ischaemia reperfusion. This study aims to investigate the effects of EGF in I/R-induced heart injury and to demonstrate its mechanisms. H9c2 cells challenged with H2O2 were used for in vitro biological activity and mechanistic studies. The malondialdehyde (MDA) and Superoxide Dismutase (SOD) levels in H9c2 cells were determined, and the cell viability was assessed by MTT assay. Myocardial I/R mouse administrated with or without EGF were used for in vivo studies. Pretreatment of H9c2 cells with EGF activated Nrf2 signalling pathway, attenuated H2O2-increased MDA and H2O2-reduced SOD level, followed by the inhibition of H2O2-induced cell death. In in vivo animal models of myocardial I/R, administration of EGF reduced infarct size and myocardial apoptosis. These data support that EGF decreases oxidative stress and attenuates myocardial ischaemia reperfusion injury via activating Nrf2.

Epidermal Growth Factor Reverses the Inhibitory Effects of the Bisphosphonate, Zoledronic Acid, on Human Oral Keratinocytes and Human Vascular Endothelial Cells In Vitro via the Epidermal Growth Factor Receptor (EGFR)/Akt/Phosphoinositide 3-Kinase (PI3K) Signaling Pathway.

BackgroundMedication-related osteonecrosis of the jaw (MRONJ) is due to the direct effects of drug toxicity and the effects on angiogenesis. The aims of this study were to evaluate the effects of treatment with the bisphosphonate, zoledronic acid, on human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs) in vitro, and whether epidermal growth factor (EGF) could alter these effects.Material/MethodsHOKs and HUVECs were incubated with zoledronic acid or EGF. Cell viability was assessed by the cell counting kit-8 (CCK-8), cell apoptosis was studied using Annexin-V conjugated to fluorescein isothiocyanate (FITC). Angiogenesis was studied by observing HUVEC tube formation and cell migrations using a transwell assay. A scratch wound assay investigated cell migration of HOKs. Western blot measured expression levels of phosphorylated epidermal growth factor receptor (EGFR), Akt, phosphoinositide 3-kinase (PI3K), the mechanistic target of rapamycin (mTOR), and endothelial nitric oxide synthase (eNOS).ResultsZoledronic acid treatment (5 μmol/L) significantly inhibited cell viability and cell migration of HOKs and HUVECs and angiogenesis of HUVECS (P<0.05); EGF partially reversed these effects (P<0.05). Zoledronic acid treatment of HOKs and HUVECs had no significant effects on apoptosis (P>0.05), but significantly reduced expression levels of p-EGFR, p-Akt, p-PI3K, p-mTOR), and p-eNOS (P<0.05); EGF partially reversed these effects and increased the expression levels (P<0.05).ConclusionsEGF partially reversed the effects of the bisphosphonate, zoledronic acid, on HOKs and HUVECs in vitro via the EGFR/Akt/PI3K signaling pathway. Further studies are required to determine the effects of EGF on MRONJ including bisphosphonate-related osteonecrosis of the jaw.

Effects of dietary supplementation with epidermal growth factor on nutrient digestibility, intestinal development and expression of nutrient transporters in early‐weaned piglets

The abnormalities in intestinal morphology and digestive function during weaning are associated with the loss of milk-borne growth factors. Epidermal growth factor (EGF) has been shown to stimulate the growth of animals. This study was to determine the effect of dietary EGF on nutrient digestibility, intestinal development and the expression of genes encoding nutrient transporters in weaned piglets. Forty-two piglets were weaned at 21days and assigned to one of three treatment groups: (1) basal diet (control), (2) basal diet+200µg/kg EGF or (3) basal diet+400µg/kg EGF. Each treatment consisted of 14 replicates, and seven piglets from each treatment were sampled on day 7 and 14. The EGF supplementation significantly elevated (p<0.05) the coefficients of total tract apparent digestibility of crude protein, calcium and phosphorus, but tended to decrease sucrase activity (p<0.10) than the control group. At day 7 post-weaning, animals receiving EGF diets showed a tendency (p<0.10) towards greater ileal villus height (VH), jejunal crypt depth (CD) and duodenal VH:CD when compared with the control group. Moreover, the mRNA levels of glucose transporter 2 (Slc2a2), neutral amino acid transporter (Slc6a19) and calbindin D9k (S100G) tended to be higher (p<0.10) for EGF groups than the control group. By day 14, EGF supplementation markedly enhanced (p<0.05) the VH, CD and VH:CD in the jejunum compared to the control group. This addition also up-regulated (p<0.05) the mRNA level and the protein abundance of peptide transporter 1 than the control group. These findings demonstrated that dietary EGF beneficially enhanced nutrient digestibility, improved intestinal development and increased the mRNA expression of nutrient transporters in weaned piglets.

Growth factor modulation of equine trophoblast mitosis and prostaglandin gene expression.

To provide insight into maternal recognition of pregnancy control in equids, the mitogenic and developmental effects of endometrium-expressed growth factors (epidermal growth factor (EGF), fibroblast growth factor 2 (FGF2), hepatocyte growth factor (HGF), and insulin-like growth factor-1 (IGF-1)) were examined in equine iTr cells, an equine trophectoderm cell line. Initial western blots revealed that HGF and IGF-1 stimulate phosphorylation of AKT serine/threonine kinase 1 (AKT1) and EGF, FGF2, or HGF resulted in phosphorylation of both extracellular signal-regulated kinase 1 (ERK1) and ERK2. Mitotic activity was stimulated (P < 0.05) by EGF, FGF2, and HGF. Chemical disruption of mitogen-activated protein kinase kinases 1 and 2 (MEK1/2) phosphorylation suppresses (P < 0.05) proliferation in control and growth factor treated cells demonstrating a dependence on ERK1/2 for mitotic activity. Treatment of iTr cells with EGF or HGF in the presence of an AKT1 inhibitor prohibits (P < 0.05) growth factor stimulated proliferation. The effect of EGF, FGF2, HGF, and IGF-1 on steroid biosynthetic enzyme gene expression, including prostaglandin-endoperoxide synthase 2 (PTGS2), was determined by real-time PCR. Neither EGF, FGF2, nor IGF-1 affected PTGS2 expression while HGF caused a two-fold increase (P < 0.05) in expression. Co-supplementation with HGF and an AKT1 inhibitor did not block PTGS2 expression, whereas providing an MEK1/2 inhibitor prevented (P < 0.05) the HGF-mediated increase in PTGS2. These results provide novel evidence of a role for HGF in equine trophectoderm proliferation and prostaglandin biosynthesis.

Inactivation of the PtdIns(4)P phosphatase Sac1 at the Golgi by H2O2 produced via Ca2+-dependent Duox in EGF-stimulated cells

Binding of epidermal growth factor (EGF) to its cell surface receptor induces production of H2O2, which serves as an intracellular messenger. We have shown that exogenous H2O2 reversibly inactivates the phosphatidylinositol 4-phosphate [PtdIns(4)P] phosphatase Sac1 (suppressor of actin 1) at the Golgi complex of mammalian cells by oxidizing its catalytic cysteine residue and thereby increases both the amount of Golgi PtdIns(4)P and the rate of protein secretion. Here we investigated the effects of EGF on Sac1 oxidation and PtdIns(4)P abundance at the Golgi in A431 cells. EGF induced a transient increase in Golgi PtdIns(4)P as well as a transient oxidation of Sac1 in a manner dependent on elevation of the intracellular Ca2+ concentration and on H2O2. Oxidation of Sac1 occurred at the Golgi, as revealed with the use of the Golgi-confined Sac1-K2A mutant. Knockdown of Duox enzymes implicated these Ca2+-dependent members of the NADPH oxidase family as the major source of H2O2 for Sac1 oxidation. Expression of a Golgi-targeted H2O2 probe revealed transient EGF-induced H2O2 production at this organelle. Our findings have thus uncovered a previously unrecognized EGF signaling pathway that links intracellular Ca2+ mobilization to events at the Golgi including Duox activation, H2O2 production, Sac1 oxidation, and PtdIns(4)P accumulation.

The Influence of TGF-β3, EGF, and BGN on SOX9 and RUNX2 Expression in Human Chondrogenic Progenitor Cells.

Osteoarthritis (OA) is the most common chronic joint disease and leads to the degradation of the extracellular matrix by an imbalance between anabolic and catabolic processes. TGF-β3 (transforming growth factor beta-3) and epidermal growth factor (EGF) influence the osteochondrogenic potential of chondrocytes. In this study, we compared the expression of mediators and receptors in the TGF-β3 and EGF pathways, as well as biglycan (BGN), in healthy and diseased chondrocytes. Furthermore, we used chondrogenic progenitor cells (CPCs) for in vitro stimulation and knockdown experiments to elucidate the effects of TGF-β3 and EGF on the chondrogenic potential. Our results demonstrate that the expression of TGF-beta receptor type-1 (TGFBRI) and epidermal growth factor receptor (EGFR) is altered in diseased chondrocytes as well as in CPCs. Moreover, TGF-β3 and EGF stimulation influenced the expression levels of BGN, SRY (sex determining region Y)-box 9 (SOX9), and Runt-related transcription factor 2 (RUNX2) in CPCs. Therefore, changes in TGFBRI and EGFR expression likely contribute to the degenerative and regenerative effects seen in late stages of OA.

Epidermal growth factor (EGF) promotes bone healing in surgically induced osteonecrosis of the femoral head (ONFH).

Angiogenic effects of epidermal growth factor (EGF), a potent mitogen, have been demonstrated previously. Moreover, different in vitro studies showed that EGF affects processes associated with bone healing, such as osteoblast differentiation and bone resorption. The aim of this study was to investigate the effect of combined core decompression (CD) and recombinant human EGF (rhEGF) treatment on early-stage osteonecrosis of the femoral head (ONFH) surgically induced in rats. ONFH was induced by dissecting the cervical periosteum and placing a ligature tightly around the femoral neck. Thirty rats were assigned to one of the following groups (n = 10 each group): sham-operated control, CD, and CD+rhEGF group. rhEGF was injected intraosseously into infarcted areas 2 weeks after the surgery. Preservation of femoral head architecture was assessed at 8 weeks post treatment by radiographic and histomorphological analyses. Osteopontin (OPN) and cluster of differentiation 31 (CD31) were detected by immunochemistry, as indicators of bone remodeling and vascular density, respectively. Inter- and intra-group (non-operated left and operated right femur) differences in radiographic and histomorphological results were analyzed. The femoral head area and sphericity were more preserved in CD+rhEGF compared to CD and sham-control group. CD31 levels were significantly different between the three groups, and were higher in CD+rhEGF compared to CD group. OPN levels were increased in CD and CD+rhEGF groups compared to sham control, but with no significant difference between CD and CD+rhEGF groups. Overall, our results indicate that EGF promotes bone formation and microvascularization in ONFH and thus positively affects the preservation of femoral head during healing.

Development of an organotypic stem cell model for the study of human embryonic palatal fusion.

Cleft palate (CP) is a common birth defect, occurring in an estimated 1 in 1,000 births worldwide. The secondary palate is formed by paired palatal shelves, consisting of a mesenchymal core with an outer layer of epithelial cells that grow toward each other, attach, and fuse. One of the mechanisms that can cause CP is failure of fusion, that is, failure to remove the epithelial seam between the palatal shelves to allow the mesenchyme confluence. Epidermal growth factor (EGF) plays an important role in palate growth and differentiation, while it may impede fusion. We developed a 3D organotypic model using human mesenchymal and epithelial stem cells to mimic human embryonic palatal shelves, and tested the effects of human EGF (hEGF) on proliferation and fusion. Spheroids were generated from human umbilical-derived mesenchymal stem cells (hMSCs) directed down an osteogenic lineage. Heterotypic spheroids, or organoids, were constructed by coating hMSC spheroids with extracellular matrix solution followed by a layer of human progenitor epithelial keratinocytes (hPEKs). Organoids were incubated in co-culture medium with or without hEGF and assessed for cell proliferation and time to fusion. Osteogenic differentiation in hMSC spheroids was highest by Day 13. hEGF delayed fusion of organoids after 12 and 18 hr of contact. hEGF increased proliferation in organoids at 4 ng/ml, and proliferation was detected in hPEKs alone. Our results show that this model of human palatal fusion appropriately mimics the morphology of the developing human palate and responds to hEGF as expected.

Potential protective effects of rhEGF against ultraviolet A irradiation-induced damages on human fibroblasts

BackgroundUltraviolet A (UVA) rays reach the dermal skin layer and generate oxidative stress, DNA damage, and cell inflammation, which in turn lead to photo-aging and photo-carcinogenesis. While there have been many studies about the beneficial effects of topical epidermal growth factor (EGF) treatment in the healing of wounds, the effect of EGF on UVA-induced skin irritation remains unknown. To clarify the effects of EGF on UVA-induced skin damage, it was investigated whether EGF signaling can affect intracellular reactive oxygen species (ROS) and DNA damages in UVA-irradiated human dermal fibroblasts.Materials and methodsFibroblasts cultured with or without rhEGF were UVA-irradiated at 40 mJ/cm2 twice per day for 5 days. After the irradiation, the intracellular ROS levels and expression of catalase and superoxide dismutase-1 (SOD-1) in the fibroblasts were ascertained. Further investigation to determine the effects of EGF on UVA-induced DNA damage, including a single cell gel electrophoresis assay and an enzyme-linked immunosorbent assay (ELISA), was carried out. Moreover, the NF-κB activity was ascertained in order to investigate the effects of EGF on UVA-irradiated fibroblasts.ResultsAs a result, it was revealed that recombinant human EGF (rhEGF) inhibited UVA- increased intracellular ROS in the fibroblasts and increased the expression of catalase and SOD-1. Moreover, in UVA-irradiated fibroblasts, the longest DNA-damaged tails were observed, but this phenomenon was not detected in cells cotreated with both UVA and rhEGF. Also, it was observed that DNA damage induction, including that of cyclobutene pyrimidine dimers, pyrimidine (6-4) pyrimidone photoproducts, and 8-hydroxy-2-deoxyguanosine, was caused by UVA irradiation. Similar to previous results, it was downregulated by rhEGF. Furthermore, rhEGF also inhibited NF-κB gene expression and the NF-κB p65 protein level in the nucleus induced by UVA irradiation.ConclusionThese results suggest that EGF might be a useful material for preventing or improving photo-aging.

Epidermal growth factor can signal via β-catenin to control proliferation of mesenchymal stem cells independently of canonical Wnt signalling

Bone marrow mesenchymal stem/stromal cells (MSCs) maintain bone homeostasis and repair through the ability to expand in response to mitotic stimuli and differentiate into skeletal lineages. Signalling mechanisms that enable precise control of MSC function remain unclear. Here we report that by initially examining differences in signalling pathway expression profiles of individual MSC clones, we identified a previously unrecognised signalling mechanism regulated by epidermal growth factor (EGF) in primary human MSCs. We demonstrate that EGF is able to activate β-catenin, a key component of the canonical Wnt signalling pathway. EGF is able to induce nuclear translocation of β-catenin in human MSCs but does not drive expression of Wnt target genes or T cell factor (TCF) activity in MSC reporter cell lines. Using an efficient Design of Experiments (DoE) statistical analysis, with different combinations and concentrations of EGF and Wnt ligands, we were able to confirm that EGF does not influence the Wnt/β-catenin pathway in MSCs. We show that the effects of EGF on MSCs are temporally regulated to initiate early “classical” EGF signalling mechanisms (e.g via mitogen activated protein kinase) with delayed activation of β-catenin. By RNA-sequencing, we identified gene sets that were exclusively regulated by the EGF/β-catenin pathway, which were distinct from classical EGF-regulated genes. However, subsets of classical EGF gene targets were significantly influenced by EGF/β-catenin activation. These signalling pathways cooperate to enable EGF-mediated proliferation of MSCs by alleviating the suppression of cell cycle pathways induced by classical EGF signalling.

Characterization of EGF-guided MDA-MB-231 cell chemotaxis in vitro using a physiological and highly sensitive assay system.

Chemotactic cell migration is a central mechanism during cancer cell invasion and hence metastasis. In order to mimic in vivo conditions, we used a three-dimensional hydrogel matrix made of collagen I and a stable gradient-generating chemotaxis assay system, which is commercially available (μ-Slide Chemotaxis) to characterize epidermal growth factor (EGF)-induced chemotaxis of the human breast cancer cell line MDA-MB-231. Surprisingly, chemotactic effects of EGF on MDA-MB-231 cells could neither be observed in the standard growth medium DMEM/F-12 supplemented with 10% serum nor in starvation medium. In contrast, after adapting the cells to the serum-free growth medium UltraCULTURETM, significant chemotactic effects could be measured with high sensitivity. The extremely time-stable linear gradients, generated in the chemotaxis chamber, led to consistent directional migration of MDA-MB-231 cells. Dose-response experiments showed increased directional and kinetic response of MDA-MB-231 cells towards stable gradients of EGF. While EGF-guided directional migration (chemotaxis) was highly concentration-dependent with the highest response at 1.5 nM/mm EGF, we found that the chemokinetic effect induced by EGF was concentration-independent. Both, blocking the ligand-binding domain of the EGF receptor by an antibody (monoclonal anti-EGFR antibody 225) and inhibition of its kinase domain by a small molecule inhibitor (AG1478) led to a reduction in EGF-induced directed migration. The high sensitivity of the assay even allowed us to observe synergistic effects in EGF-receptor inhibition using a combination of low doses of both inhibitor types. Those results validate the fact that EGF is a potent guidance cue for MDA-MB-231 cell migration and help to understand the mechanism behind chemotaxis-driven cancer metastasis.

Topical administration of EGF suppresses immune response and protects skin barrier in DNCB-induced atopic dermatitis in NC/Nga mice

Atopic dermatitis (AD) is a common inflammatory skin disease characterized by a complex, heterogeneous pathogenesis including skin barrier dysfunction, immunology, and pruritus. Although epidermal growth factor (EGF) is essential for epithelial homeostasis and wound healing, the effect of EGF on AD remains to be explored. To develop a new therapy for AD, the anti-AD potential of EGF was investigated by inducing AD-like skin lesions in NC/Nga mice using 2,4-dinitrochlorobenzene (DNCB). EGF was administrated to NC/Nga mice to evaluate its therapeutic effect on DNCB-induced AD. EGF treatment improved dermatitis score, ear thickness, epidermal hyperplasia, serum total immunoglobulin E level, and transepidermal water loss in NC/Nga mice with DNCB-induced AD. In addition, levels of skin barrier-related proteins such as filaggrin, involucrin, loricrin, occludin, and zonula occludens-1 (ZO-1) were increased by EGF treatment. These beneficial effects of EGF on AD may be mediated by EGF regulation of Th1/Th2-mediated cytokines, mast cell hyperplasia, and protease activated receptor-2 (PAR-2) and thymic stromal lymphopoietin (TSLP), which are triggers of AD. Taken together, our findings suggest that EGF may potentially protect against AD lesional skin via regulation of skin barrier function and immune response.

Novel role of Giα2 in cell migration: Downstream of PI3-kinase-AKT and Rac1 in prostate cancer cells.

Tumor cell motility is the essential step in cancer metastasis. Previously, we showed that oxytocin and epidermal growth factor (EGF) effects on cell migration in prostate cancer cells require Giα2 protein. In the current study, we investigated the interactions among G-protein coupled receptor (GPCR), Giα2, PI3-kinase, and Rac1 activation in the induction of migratory and invasive behavior by diverse stimuli. Knockdown and knockout of endogenous Giα2 in PC3 cells resulted in attenuation of transforming growth factor β1 (TGFβ1), oxytocin, SDF-1α, and EGF effects on cell migration and invasion. In addition, knockdown of Giα2 in E006AA cells attenuated cell migration and overexpression of Giα2 in LNCaP cells caused significant increase in basal and EGF-stimulated cell migration. Pretreatment of PC3 cells with Pertussis toxin resulted in attenuation of TGFβ1- and oxytocin-induced migratory behavior and PI3-kinase activation without affecting EGF-induced PI3-kinase activation and cell migration. Basal- and EGF-induced activation of Rac1 in PC3 and DU145 cells were not affected in cells after Giα2 knockdown. On the other hand, Giα2 knockdown abolished the migratory capability of PC3 cells overexpressing constitutively active Rac1. The knockdown or knockout of Giα2 resulted in impaired formation of lamellipodia at the leading edge of the migrating cells. We conclude that Giα2 protein acts at two different levels which are both dependent and independent of GPCR signaling to induce cell migration and invasion in prostate cancer cells and its action is downstream of PI3-kinase-AKT-Rac1 axis.

The effects of recombinant human epidermal growth factor containing ointment on wound healing and post inflammatory hyperpigmentation prevention after fractional ablative skin resurfacing: A split-face randomized controlled study.

Epidermal growth factor (EGF) is one of the important peptides in wound healing process. The effects of EGF have been increasingly studied in various types of ulcers. However, data on postablative laser resurfacing wound is still limited. To evaluate the effects of the topical EGF ointment on wound healing process and postinflammatory hyperpigmentation (PIH) prevention after fractional ablative laser resurfacing. This is a randomized split-face study. Nineteen healthy subjects were enrolled and completed follow up protocol. Patients received single treatment of fractional carbon dioxide laser on both cheeks. After randomization, each patient was assigned to apply one side of the face with topical EGF ointment and another side with petrolatum. Wound healing was evaluated by duration of scab shedding, duration of postlaser erythema, erythema index, and transepidermal water loss on the daily follow up period of seven days after treatment. PIH was evaluated at 2, 3weeks and 1, 2months follow up by photographs and melanin index. Most of patients were female with Fitzpatrick skin phototype III to V. Comparing with control (petrolatum), EGF treated side showed no significant difference in duration of scab shedding, duration of postlaser erythema, erythema index, and transepidermal water loss (P-value=.58, .22, .78, and .51, respectively). Incidence of PIH was 52.6% on EGF side and 57.9% on petrolatum side, however, it was not statistically different (P=.56). The melanin index was also not different as well (P=.96). Topical EGF might provide significant wound healing stimulation for chronic wound more than acute wound. Further studies, especially in post laser wound or other cosmetic purposes are needed.

Cyclic Guanosine Monophosphate (cGMP)-Dependent Protein Kinase II Blocks Epidermal Growth Factor (EGF)/Epidermal Growth Factor Receptor (EGFR)-Induced Biological Effects on Osteosarcoma Cells.

BackgroundThe present work was performed to detect the potential inhibitory effect of cyclic guanosine monophosphate (cGMP)-dependent protein kinase II (PKG II) on epidermal growth factor (EGF) receptor-induced biological activity and related signal cascades in osteosarcoma cells.Material/MethodsWe transfected the osteosarcoma MG-63 cell line with an adenoviral vector encoding PKG II cDNA (Ad-PKGII) and incubated the transfected cells with 250 μM 8-pCPT-cGMP to activate the PKG II. We stimulated the MG-63 cells with100 ng/ml EGF, and then detected their proliferation using a CCK-8 assay. Transwell assay was used to examine MG-63 cell migration; and Western blot analysis was used to detect expression of matrix metalloproteinase 9 (MMP-9) and activation of ERK and Akt.ResultsStimulating cells by 100 ng/ml EGF promoted MG-63 cell proliferation and migration, ERK and Akt phosphorylation, and MMP-9 expression. These effects of EGF were inhibited in MG-63 cells infected with Ad-PKGII and incubated with 8-pCPT-cGMP.ConclusionsOur results demonstrate that Ad-PKGII infection significantly inhibited EGF-induced proliferation and migration, as well as the associated-signal cascades; which indicates that PKG II might be a potential anti-cancer factor.

Epidermal Growth Factor, through Alleviating Oxidative Stress, Protect IPEC-J2 Cells from Lipopolysaccharides-Induced Apoptosis.

The epidermal growth factor (EGF) has been widely used for protection of stress-induced intestinal mucosa dysfunction. However, whether EGF would alleviate oxidative injury and reduce apoptosis in porcine intestine is not yet known. Therefore, the aim of this study was to investigate the effect of EGF on lipopolysaccharides (LPS)-induced induction of oxidative stress and ensuing apoptosis in the porcine intestinal epithelial cell line, IPEC-J2. The present study showed that EGF significantly increased cell viability and decreased the LPS-induced induction of apoptosis, dehydrogenase (LDH) release and malonaldehyde (MDA) production. EGF also (i) decreased expression of the pro-apoptotic genes Fas, Bax, Cascase-3, Cascase-8, Cascase-9, and proteins such as P53, Fas, Bax, Caspase3; (ii) increased antiapoptotic protein B-cell lymphoma 2 (Bcl2) expression; (iii) increased mRNA levels of the nuclear factor erythroid 2-related factor 2 (Nrf2) related genes Nrf2, manganese superoxide dismutase (SOD2), catalase (CAT), glutathione peroxidase (GSH-Px), heme oxygenase (HO-1) and quinone oxidoreductase (NQO1); (iv) protein level of Nrf2-realeted proteins Nrf2, HO-1, NQO1; and (v) total antioxidant capacity (T-AOC), CAT, SOD, GSH-Px concentrations. Collectively, our results indicated that EGF enhanced Nrf2 protein expression, and upregulated the expression of phase II metabolizing enzymes (such as HO-1 and NQO1) and antioxidative enzymes (SOD, CAT and GSH-Px) to alleviate oxidative injury, and then protect IPEC-J2 cells from apoptosis induced by LPS.

Entrainment of the Circadian Clock in Neural Stem Cells by Epidermal Growth Factor is Closely Associated with ERK1/2-mediated Induction of Multiple Clock-related Genes

The mitotic activity of certain tissues in the body is closely associated with circadian clock function. However, the effects of growth factors on the molecular clockwork are not fully understood. Stimulation of neural stem cells (NSCs) with epidermal growth factor (EGF), a well-known mitogen, is known to cause synchronized cell cycle progression with a period of approximately 24 h, closely associated with the Per2 gene expression rhythm. Here, we examined the effects of EGF on the molecular clockwork of NSCs. Treatment of cultured NSCs derived from embryonic mouse forebrain with EGF (20 ng/mL) caused a phase shift in the PER2::LUCIFERASE bioluminescence rhythm in a stimulation time-dependent manner. The EGF phase-response curve differed from that of forskolin (FK)—a well-known chemical resetting stimulus—both in the advance/delay ratio and stimulation time-dependency. PCR array analysis followed by quantitative PCR validation demonstrated that EGF treatment transiently induced multiple clock-related genes including Per1, Per2, Dec1, e4bp4, and Noct, whereas FK treatment induced a limited number of genes (Per1 and Dec1), suggesting that the mode of entrainment of NSC molecular clock was different for EGF and FK. EGF led to gene induction in the presence of cycloheximide, suggesting that de novo protein synthesis is unnecessary. Pretreatment with the MEK1/2 inhibitor U0126 significantly suppressed the acute induction of Per2, Dec1, and Noct by EGF and also abolished the EGF-induced phase shift of the PER2::LUCIFERASE rhythm in NSCs. These results suggest a unique effect of EGF on the molecular clockwork of NSCs.

Epidermal Growth Factor Attenuated the Expression of Inflammatory Cytokines in Human Epidermal Keratinocyte Exposed to Propionibacterium acnes.

BackgroundWhile the beneficial effects of topical epidermal growth factor (EGF) on wound healing have been repeatedly reported, there are few reports about the effects of EGF on inflammatory skin diseases including acne.ObjectiveTo clarify the effects of EGF on acne, it was investigated whether recombinant human EGF (rhEGF) signalling can affect Propionibacterium acnes-induced cytokine expression in human epidermal keratinocytes.MethodsThe cultured normal human epidermal keratinocytes (NHK) were co-treated with P. acnes and rhEGF, and mRNA levels of interleukin (IL)-1α, IL-8 and tumor necrosis factor (TNF)-α; toll-like receptor 2 (TLR2); and nuclear factor kappa B (NF-κB) were determined. Specific enzyme-linked immunosorbent assay kits were used to measure the IL-1α, IL-8 and TLR2 expression as well as the NF-κB activation in P. acnes and rhEGF-treated NHK. After infecting the cultured NHK with live P. acnes, an increased expression of IL-1α, IL-8 and TNF-α was detected, which was prevented by rhEGF co-treatment.ResultsTLR2 and NF-κB activity increased after P. acnes treatment, and rhEGF treatment decreased TLR2 expression and NF-κB activity dose-dependently. The inhibition of EGF receptor by gefitinib attenuated the inhibitory effect of rhEGF on these increased expressions of proinflammatory cytokines and TLR2 and activity of NF-κB in NHK stimulated by P. acnes.ConclusionThese results suggest that EGF attenuated P. acnes-induced inflammatory responses, at least in part, through the modulation of TLR2 signalling, and the topical application of rhEGF may be beneficial to relieve the inflammatory reactions of acne.