Heparin‐modified dendrimer crosslinked collagen matrices for the delivery of heparin‐binding epidermal growth factor

Heparin-Binding Epidermal Growth Factor–Like Growth Factor as a Critical Mediator of Tissue Repair and Regeneration

a critical issue in understanding how the mammalian kidney responds to pathological states and nephrotoxic chemicals involves the identification of modulating agents, growth factors, and signaling pathways that regulate responses. Although some of the modulating agents identified, including

BLADDER EPITHELIAL CELLS FROM PATIENTS WITH INTERSTITIAL CYSTITIS PRODUCE AN INHIBITOR OF HEPARIN-BINDING EPIDERMAL GROWTH FACTOR-LIKE GROWTH FACTOR PRODUCTION

Role of membrane-bound heparin-binding epidermal growth factor-like growth factor (HB-EGF) in renal epithelial cell branching

Deletion of the epidermal growth factor receptor in renal proximal tubule epithelial cells delays recovery from acute kidney injury

Keratinocyte Growth Inhibition by High-Dose Epidermal Growth Factor Is Mediated by Transforming Growth Factor β Autoinduction: A Negative Feedback Mechanism for Keratinocyte Growth

We have analyzed ErbB receptor interplay induced by the epidermal growth factor (EGF)-related peptides in cell lines naturally expressing the four ErbB receptors. Down-regulation of cell surface ErbB-1 or ErbB-2 by intracellular expression of specific antibodies has allowed us to delineate the role of these receptors during signaling elicited by: EGF and heparin binding EGF (HB-EGF), ligands of ErbB-1; betacellulin (BTC), a ligand of ErbB-1 and ErbB-4; and neu differentiation factor (NDF), a ligand of ErbB-3 and ErbB-4. Ligand-induced ErbB receptor heterodimerization follows a strict hierarchy and ErbB-2 is the preferred heterodimerization partner of all ErbB proteins. NDF-activated ErbB-3 or ErbB-4 heterodimerize with ErbB-1 only when no ErbB-2 is available. If all ErbB receptors are present, NDF receptors preferentially dimerize with ErbB-2. Furthermore, EGF- and BTC-induced activation of ErbB-3 is impaired in the absence of ErbB-2, suggesting that ErbB-2 has a role in the lateral transmission of signals between other ErbB receptors. Finally, ErbB-1 activated by all EGF-related peptides (EGF, HB-EGF, BTC and NDF) couples to SHC, whereas only ErbB-1 activated by its own ligands associates with and phosphorylates Cbl. These results provide the first biochemical evidence that a given ErbB receptor has distinct signaling properties depending on its dimerization.

Adiponectin Deficiency Protects Mice From Chemically Induced Colonic Inflammation

Epidermal growth factor receptor (EGFR) has been implicated in tumour growth and extension of ovarian cancer. Peritoneal fluid in ovarian cancer patients contains various growth factors that can promote tumour growth and extension. In order to investigate the clinical significance of EGFR ligands as activating factors of ovarian cancer, we examined the cell proliferation-promoting activity and the level of EGFR ligands in peritoneal fluid obtained from 99 patients. Proliferation-promoting activity in peritoneal fluid from 63 ovarian cancer patients (OVCA) was much higher than peritoneal fluid from 18 ovarian cyst patients (OVC) and 18 normal ovary patients (NO), and the activity was suppressed only by antibodies against EGFR or heparin-binding epidermal growth factor (HB-EGF). A large difference was observed in the level of EGFR ligands between HB-EGF and TGF-α or amphiregulin. The concentration of HB-EGF in OVCA significantly increased compared to that in OVC or NO (P<0.01). No significant difference in the concentration of TGF-α and amphiregulin was found between the OVCA and NO or OVC groups. In peritoneal fluid, HB-EGF is sufficiently elevated to activate cancer cells even at an early stage of OVCA. These results suggested that HB-EGF in peritoneal fluid might play a key role in cell survival and in the proliferation of OVCA.

Diminished survival of human cytotrophoblast cells exposed to hypoxia/reoxygenation injury and associated reduction of heparin-binding epidermal growth factor-like growth factor

The mechanism behind the growth-promoting effect of insulin is a subject of debate. Employing RT4 bladder cancer cells, we examined the cross-talk between insulin and the epidermal growth factor system. We found that insulin induced a time- and dose-dependent (25-1000 nmol.L(-1) insulin) increase in mRNA expression of three ligands from the epidermal growth factor system. Times for peak increase and fold increase after incubation with 250 nmol.L(-1) insulin were as follows: heparin-binding epidermal growth factor-like growth factor, 0.5 h, 1.4-fold, P < 0.05; epiregulin, 3 h, 14-fold, P < 0.0001; and amphiregulin, 3 h, 12-fold, P < 0.001. Induction of heparin-binding epidermal growth factor-like growth factor and amphiregulin was verified at the protein level. We demonstrate that incubation of RT4 bladder cancer cells for 24 h with 250 nmol.L(-1) insulin increases proliferation by 43% (P < 0.0001) as compared to untreated cells. At the same time, phosphorylation and thereby activation of the epidermal growth factor receptor (HER1) was observed. Both phosphorylation and insulin-induced proliferation were almost completely inhibited by the HER1 inhibitor Iressa (P < 0.0001). This shows that insulin leads to activation of HER1, and that HER1 plays an essential role in mediating the growth-promoting effect of insulin. Iressa inhibited not only the activation of HER1 caused by insulin but also the insulin-induced increase in the three ligands (heparin-binding epidermal growth factor-like growth factor, epiregulin and amphiregulin). As heparin-binding epidermal growth factor-like growth factor was induced before epiregulin and amphiregulin upon insulin stimulation, we speculated that the insulin-induced heparin-binding epidermal growth factor-like growth factor initiated the activation of HER1, and that this in turn led to increased expression of epiregulin and amphiregulin and thereby to continued activation of HER1. Earlier reports have shown that insulin-like growth factor receptor can activate HER1 via its ligand heparin-binding epidermal growth factor-like growth factor. In accord with this, we found that treatment of RT4 cells with recombinant heparin-binding epidermal growth factor-like growth factor mimicked the effect of insulin, with induction of mRNA for the three ligands. However, the insulin-induced increase in mRNA expression of amphiregulin and epiregulin could not be prevented by the heparin-binding epidermal growth factor-like growth factor inhibitor CRM197, demonstrating that heparin-binding epidermal growth factor-like growth factor is not essential for the insulin-induced increase in the expression of these ligands. In conclusion, we show that insulin-induced growth in RT4 cells requires activated HER1. Furthermore, activation of HER1 is required for the insulin-induced increase in expression of the HER1 ligands heparin-binding epidermal growth factor-like growth factor, amphiregulin and epiregulin.

Up-regulation of heparin binding epidermal growth factor-like growth factor and amphiregulin expression in Helicobacter pylori-infected human gastric mucosa

Several growth factors including hepatocyte growth factor (HGF) have been implicated in the regulation of liver regeneration. Recently, we reported that heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF) has hepatotrophic effects in vitro. We investigated the role of HB-EGF as a hepatotrophic factor in regenerating rat liver after 70% partial hepatectomy. The level of HB-EGF messenger RNA (mRNA) in regenerating rat liver increased 1.5 hours after partial hepatectomy and reached a maximum (about sevenfold over normal) at 6 hours. In contrast, hepatic HGF mRNA levels increased at 12 hours and achieved maximal expression at 24 hours. HB-EGF protein expression increased about 2.8-fold over normal at 10 hours after partial hepatectomy. The number of EGF receptors, to which HB-EGF binds, decreased 6 hours after partial hepatectomy. HB-EGF mRNA levels increased in nonparenchymal cells (NPCs) at 6 hours after partial hepatectomy but not in hepatocytes. Using the reverse transcription-polymerase chain reaction (RT-PCR), HB-EGF gene expression was increased predominantly in Kupffer cells and sinusoidal endothelial cells but not in lipocytes and hepatocytes. These results indicated that HB-EGF may be an important growth factor, produced in an earlier phase rather than HGF, in the regenerating liver after partial hepatectomy by a paracrine mechanism.

The dysregulation of EGF family ligand cleavage has severe consequences for the developing as well as the adult organism. Therefore, their production is highly regulated. The limiting step is the ectodomain cleavage of membrane-bound precursors by one of several a disintegrin and metalloprotease (ADAM) metalloproteases, and understanding the regulation of cleavage is an important goal of current research. We have previously reported that in mouse lung epithelial cells, the pro-EGF ligands TGFα, neuregulin 1β (NRG), and heparin-binding EGF are differentially cleaved depending on the cleavage stimulus (Herrlich, A., Klinman, E., Fu, J., Sadegh, C., and Lodish, H. (2008) FASEB J.). In this study in mouse embryonic fibroblasts that lack different ADAMs, we show that induced cleavage of EGF ligands can involve the same substrate-specific metalloprotease but does require different stimulus-dependent signaling pathways. Cleavage was stimulated by phorbol ester (12-O-tetradecanoylphorbol-13-acetate (TPA), a mimic of diacylglycerol and PKC activator), hypertonic stress, lysophosphatidic acid (LPA)-induced G protein-coupled receptor activation, or by ionomycin-induced intracellular calcium release. Although ADAMs showed substrate preference (ADAM17, TGFα and heparin-binding EGF; and ADAM9, NRG), substrate cleavage differed substantially with the stimulus, and cleavage of the same substrate depended on the presence of different, sometimes multiple, PKC isoforms. For instance, classical PKC was required for TPA-induced but not hypertonic stress-induced cleavage of all EGF family ligands. Inhibition of PKCζ enhanced NRG release upon TPA stimulation, but it blocked NRG release in response to hypertonic stress. Our results suggest a model in which substantial regulation of ectodomain cleavage occurs not only on the metalloprotease level but also on the level of the substrate or of a third protein.

Epidermal growth factor (EGF) plays an important role in corneal epithelial migration and proliferation to improve the wound healing process. This study aimed to understand the role of NFκB in EGF-induced corneal epithelial wound healing through regulation of CTCF activity, which plays important roles in cell motility and migration to promote wound healing. The effect of NFκB p50 on corneal epithelial wound healing was investigated by comparing the eyes of wild-type and p50 knockout mice. We found that there was a significant retardation in corneal epithelial wound healing in the corneas of p50 knockout mice. Wound closure rates were measured in human corneal epithelial cells transfected with an NFκB activation-sensitive CTCF expression construct to demonstrate the effect of human CTCF expression under the control of EGF-induced NFκB activation on wound healing. EGF stimulation activated NFκB, which directly triggered the expression of the exogenous human CTCF in transfected cells and, subsequently, promoted human corneal epithelial cell motility, migration, and wound healing. Overexpression of CTCF in corneal epithelial cells and mouse corneas significantly enhanced the wound healing process. Furthermore, the effect of overexpressing NFκB p50 in corneal epithelial cells on the promotion of wound healing was abolished by knockdown of CTCF with CTCF-specific shRNA. Thus, a direct regulatory relationship between EGF-induced NFκB p50 and CTCF activation affecting corneal epithelial wound healing has been established, indicating that CTCF is, indeed, a NFκB p50-targeted and effective gene product in the core transcriptional network downstream from the growth factor-induced NFκB signaling pathway.