Background and Aims: Several single nucleotide polymorphisms (SNPs), including rs4444903 in the epidermal growth factor (EGF) gene, rs12979860 near the interleukin-28B (IL28B) gene, and rs738409 in the patatin-like phospholipase domain-containing protein 3 (PNPLA3) gene, have been linked to treatment response, steatosis, fibrosis, and development of hepatocellular carcinoma (HCC) in chronic hepatitis C (HCV). No study has comprehensively examined the effects of these SNPs on the natural history of HCV-related cirrhosis. Methods: We performed a retrospective cohort study of 169 subjects with chronic HCV and biopsy-proven cirrhosis who had long term followup for clinical events. Formalin-fixed, paraffin-embedded liver biopsy specimens were genotyped for EGF, IL28B, and PNPLA3 using a TaqMan assay with commercial probes and primers. Cox proportional hazards modeling was used to determine the hazard ratio for clinical decompensation, defined as the development of ascites, encephalopathy, variceal hemorrhage, HCC, or cirrhosis-related death. Results: During a median followup of 6.6 years, 66 patients (39%) experienced clinical decompensation. On univariate analysis, EGF non-A/A, PNPLA3 non-C/C, and IL28B non-C/C genotypes were each associated with increased risk of decompensation. In multivariable Cox regression modeling, EGF non-A/A genotype was independently associated with an increased rate of clinical decompensation (HR = 3.00, p = 0.005). Conclusions: HCV cirrhotics with the EGF A/G and G/G genotypes at rs4444903, a functional polymorphism associated with higher intrahepatic EGF levels, have an increased risk of clinical decompensation. Further study of the predictive value of EGF genotyping in patients with earlier stages and other etiologies of liver disease is warranted.

Epidermal Growth Factor (EGF) is a member of the EGF-like family. EGF binding to the Epidermal Growth Factor Receptor (EGFR) affects cell survival as well as proliferation, migration, and tissue differentiation. Over-expression along with ligand-induced activation of EGFR has been correlated with increased in vivo invasiveness of tumor cells and enhanced in vitro migration of cell lines. This in turn makes EGFR a very important target for cancer therapy. In our investigation we have discovered that the cytoplasmic domain of proEGF (proEGFcyt) has the ability to decrease proliferation and migration in thyroid carcinoma cell lines. We illustrated that proEGFcyt causes specific alterations in the microtubular (MT) phenotype and the composition of MT-associated proteins (MAP) in FTC-133 overexpressing proEGFcyt, a finding not observed in FTC-133 over-expressing a novel splice form of proEGFcyt with a deletion of the complete exon 23 (proEGFdel23) (Pyka et al., 2004). Here, we demonstrate that proEGFcyt suppresses motility and elastinolytic activity in human thyroid Ca cells as a result from reduced secretion of cath-L. This impaired the ability of thyroid Ca cells to penetrate elastin matrices. The reduction in cath-L secretion was as a result of an up-regulation of SNAP25, a member of the t-SNARE plasma membrane complex, which is involved in Ca2+-dependent exocytosis. Furthermore, we demonstrated that proEGFcyt-mediated silencing of UCH-L1 causes the decrease in EGFR due to enhanced EGFR ubiquitination. This correlated with altered proteasomal degradation and provides a unique new mechanism on how proEGFcyt can affect cell proliferation in human thyroid cancer cells. These studies identified novel functions of human proEGF as a bidirectional signaling molecule consisting of the known extracellular EGF domain which functions as the classical ligand and activator of EGFR-mediated cell growth and the proEGF cytoplasmic domain which has the ability to suppress migration and growth of human thyroid carcinoma cells.

Epidermal Growth Factor (EGF) is a member of the EGF-like family. EGF binding to the Epidermal Growth Factor Receptor (EGFR) affects cell survival as well as proliferation, migration, and tissue differentiation. Over-expression along with ligand-induced activation of EGFR has been correlated with increased in vivo invasiveness of tumor cells and enhanced in vitro migration of cell lines. This in turn makes EGFR a very important target for cancer therapy. In our investigation we have discovered that the cytoplasmic domain of proEGF (proEGFcyt) has the ability to decrease proliferation and migration in thyroid carcinoma cell lines. We illustrated that proEGFcyt causes specific alterations in the microtubular (MT) phenotype and the composition of MT-associated proteins (MAP) in FTC-133 overexpressing proEGFcyt, a finding not observed in FTC-133 over-expressing a novel splice form of proEGFcyt with a deletion of the complete exon 23 (proEGFdel23) (Pyka et al., 2004). Here, we demonstrate that proEGFcyt suppresses motility and elastinolytic activity in human thyroid Ca cells as a result from reduced secretion of cath-L. This impaired the ability of thyroid Ca cells to penetrate elastin matrices. The reduction in cath-L secretion was as a result of an up-regulation of SNAP25, a member of the t-SNARE plasma membrane complex, which is involved in Ca2+-dependent exocytosis. Furthermore, we demonstrated that proEGFcyt-mediated silencing of UCH-L1 causes the decrease in EGFR due to enhanced EGFR ubiquitination. This correlated with altered proteasomal degradation and provides a unique new mechanism on how proEGFcyt can affect cell proliferation in human thyroid cancer cells. These studies identified novel functions of human proEGF as a bidirectional signaling molecule consisting of the known extracellular EGF domain which functions as the classical ligand and activator of EGFR-mediated cell growth and the proEGF cytoplasmic domain which has the ability to suppress migration and growth of human thyroid carcinoma cells.

No description available.

In the U.K. deaths due to liver diseases, especially alcohol related diseases, have risen considerably over the last 20 years. In 2005 up to 13,000 people died from liver related diseases within the U.K., including alcohol and viral liver failure and liver cancers. Worldwide hepatitis B affects about 2 billion people, killing 500,000 to 1 million per year. An effective way to treat liver disease is often liver surgery, such as liver resection for cancers and liver transplant for failure. However, the failure of liver regeneration by hepatocyte proliferation after resection surgery leads to a high death rate, and a shortage of liver donors means most people with liver failure die without access to a transplant. Therefore, understanding hepatocyte proliferation is a key to improving survival after resection surgery and providing hepatocytes for cell therapy in place of organ donation. The mechanism of hepatocyte proliferation has been studied both in vivo and in culture by many groups. However, only limited proliferation and preservation of function of primary human and rat hepatocytes, not suitable for clinical use, has been achieved on stimulation with growth factors. This study focuses on the mechanism of epidermal growth factor (EGF) stimulation of rat hepatocyte cell cycle progression and proliferation, including the role of PI3K/Akt/mTOR and MEK/ERK signalling pathways, EGF receptor location after activation of downstream proteins such as protein kinase B (Akt) and extracellular signal-regulated kinases 1/2 (ERK1/2), and their effect on the cell cycle. Included in this study are some comparisons between the stimulation of the EGF receptor (a tyrosine kinase receptor) and the P2Y receptor (a G protein coupled receptor). The PI3K/Akt/mTOR signalling pathway appears to be necessary for the hepatocyte response to EGF, inducing progression to S phase and DNA synthesis, while the MEK/ERK pathway is important but not necessary. The P2Y2 agonist UTP, which also stimulates these two pathways, does not result in the cell entering S phase. This suggests that the activation of these two signalling pathways is not sufficient for cell cycle progression. Furthermore, infection of cells with adenovirus to express constitutively active Akt increases hepatocytes proliferation and induces cell cycle progression, which generates a window to obtain hepatocytes proliferation in culture. It has been shown in this thesis that EGF stimulation of ERK phosphorylation continues from endosomes, while the evidence suggests that UTP stimulation is restricted to signalling at the cell surface. Furthermore, endocytic EGF/EGFR alone (without stimulation from the cell surface) is sufficient to induce cell cycle progression. This endosomal signalling with EGF but not UTP may explain the absence of cell cycle progression following UTP. EGF stimulates the appearance of phospho-EGFR in the nucleus. Furthermore, nuclear EGFR has a different apparent molecule weight than the cytoplasmic receptor; this may be due to nuclear EGFR having fewer and/or different phosphates. In vivo work by others has shown that in liver regeneration following partial hepatectomy (PH) EGF and full-length activated-EGFR were showed to be present in proliferating hepatocytes. This thesis describes the mechanism of growth factor (EGF) stimulation of primary rat hepatocyte proliferation. It shows for the first time that endosomal EGF/EGFR alone is sufficient to stimulate cell cycle progression, and that EGF induces phospho-EGFR in the nucleus in cultured rat hepatocytes. This thesis also provides the possibility to obtain cultured hepatocytes proliferation including over-expression of constitutively active form of Akt and translocation to the nucleus of full-length EGFR in the phosphorylated form. These studies improve our understanding of growth factor (e.g. EGF) stimulation of hepatocyte proliferation in vitro and help to move closer to the goal of obtaining sufficient functional hepatocytes in culture for clinical use, and of drugs that will stimulate hepatocyte proliferation following resection surgery.

The structure and function of brain shows sexual dimorphism in the vertebrates. Sexual differentiation is divided into brain sexual differentiation and gonad sexual differentiation. Brain sexual differentiation is resulted from the neural development. In the present study, the neurosphere cloned from tilapia, Oreochromis mossambicus, was used. The characteristics of neurosphere derived from both primary- and sub- culture, were studied. The effects of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF, FGF2), and temperature on the neurosphere cloned from both primary culture and subculture, were investigated. These results show that the neurospheres, cloned form both primary- and sub- culture, is consist of the nestin-immunoreactive cell. Furthermore, the cell of the neurosphere shows an ability of differentiation. And the diameter of neurosphere in the subculture is significantly larger than that of primary culture. On the other hand, both FGF and temperature have an effect to increase the diameter of neurosphere in the primary culture.

Abstract Integrin-linked Kinase (ILK) is an integrin-associated protein kinase, which regulates growth factor-signalling pathways and cell-ECM adhesion events. Abrogated ILK expression or activity has been implicated in contributing to oncogenic transformation. We examined the role played by ILK in growth factor-stimulated and integrin signalling events in five human oesophageal squamous cell carcinoma cell lines (HOSCCs), known to overexpress the EGF receptor. Western blot analysis revealed the presence of ILK (59kDa) in all the moderately differentiated HOSCC lines. ILK1 was confirmed as being the predominant isoform. Densitometrically analysed Western blots showed that, per unit of protein, ILK is expressed uniformly across the cell lines under standard culture conditions. Following EGF (10 ng/ml) and TGFβ1 (1 ng/ml) treatment, ILK expression increased in all five HOSCCs. Indirect immunofluorescence microscopy showed the majority of ILK to localise at a cytoplasmic/nuclear level, with a proportion of ILK localising at the membrane, which resembled the distribution pattern of the β3 integrin subunit. This membranal distribution most likely follows that of the adhesion plaques although lesser, and variable, amounts were also identified throughout the cytoplasm. The functionality of the ILK1 kinase domain was demonstrated using myelin basic protein (MBP)-based kinase assays. EGF and TGFβ1 treatment produced an increase in ILK activity in the WHCO3 cell line of 3.5 fold, but a decrease in activity in the other cell lines, which are suggested to involve the actions of PTEN. The identification of nuclear ILK was surprising, and the mechanism for nuclear ILK localisation was suggested to involve a caveolae-associated protein, caveolin-1. Cell adhesion assays revealed that KP-392-mediated inhibition of ILK resulted in a nonsignificant reduction in cell adhesion to collagen and fibronectin. These data provide distinctive evidence for the influence of growth factors on ILK expression, but a duality in the effect on ILK activity. This apparent dichotomy is noteworthy and may be of particular relevance in HOSCC. It is further suggested that KP-392-induced ILK inhibition destabilises the αβ integrin heterodimer and that PI3K acts upstream of ILK-mediated cell adhesion events in HOSCCs. This suggests that ILK mediates integrin associated processes in human oesophageal SCC cell lines.

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No description available.

No description available.