Conditional knockout of heparin‐binding epidermal growth factor‐like growth factor in the liver accelerates carbon tetrachloride‐induced liver injury in mice

Aim: We previously demonstrated that heparin‐binding epidermal growth factor‐like growth factor (HB‐EGF) is induced in response to several liver injuries. Because the HB‐EGF knockout (KO) mice die in utero or immediately after birth due to cardiac defects, the loss of function study in vivo is limited. Here, we generated liver‐specific HB‐EGF conditional knockout mice using the interferon‐inducible Mx‐1 promoter driven cre recombinase transgene and investigated its role during acute liver injury. Methods: We induced acute liver injury by a single i.p. injection of carbon tetrachloride (CCl₄) in HB‐EGF KO mice and wild‐type mice and liver damage was assessed by biochemical and immunohistochemical analysis. We also used AML12 mouse hepatocyte cell lines to examine the molecular mechanism of HB‐EGF‐dependent anti‐apoptosis and wound‐healing process of the liver in vitro. Results: HB‐EGF KO mice exhibited a significant increase of alanine aminotransferase level and also showed a significant increase in the number of apoptotic hepatocytes assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling staining at 24 h after CCl₄ injection. We also demonstrated that HB‐EGF treatment inhibited tumor necrosis factor‐α‐induced apoptosis of AML12 mouse hepatocytes and promoted the wound‐healing response of these cells. Conclusion: This study showed that HB‐EGF plays a protective role during acute liver injury.     

New mechanism of transforming growth factor‐β signaling in hepatoma: Dramatic up‐regulation of tumor initiating cells and epidermal growth factor receptor expression

Aim: Transforming growth factor‐β (TGF‐β) has dual activity in tumor cells. We studied the effect of TGF‐β on tumor‐initiating cells (TICs), which are similar in self‐renewal and differentiation features to normal adult stem cells. Methods: We used side population (SP) cells that exclude DNA binding dye Hoechst 33342 to obtain TICs, studied the differences in the kinetics of the SP cell response to TGF‐β treatment between hepatic tumor cell lines, and performed gene analysis. Results: SP cells from all cell lines have higher proliferative ability compared to non‐SP cells and they are drug resistant. TGF‐β treatment increased the percentage of SP cells (%SP) and the survival rate; chemotherapeutic drug resistance developed only in K‐251 SP cells. Gene analysis showed that TGF‐β up‐regulated epidermal growth factor receptor (EGFR) only in K‐251 cells. There were no EGFR mutations in K‐251, which had been reported in lung cancer. Knockdown of Smad4 using the small‐interfering RNA technique in K‐251 cells inhibited EGFR overexpression and significantly decreased the %SP. In contrast, the JNK inhibitor had little effect on EGFR expression or the %SP. Conclusion: TGF‐β treatment of K‐251 cells causes tumor progression and the anti‐cancer drug resistant phenotype by increasing SP.     

HCV core protein promotes heparin binding EGF‐like growth factor expression and activates Akt

Aims: Persistent hepatitis C virus (HCV) infection is a major cause of chronic liver dysfunction and is closely associated with the development of human hepatocellular carcinoma (HCC). Among HCV components, core protein is implicated in cell growth regulation, and we previously demonstrated that HCV core protein interacted with 14‐3‐3 protein and activated the kinase Raf‐1 and mitogen‐activated protein kinase (MAPK)/extracellular regulated kinase (ERK) pathway. In the present study, we investigated the expression levels and function of downstream molecules in the MAPK/ERK signaling pathway in cells expressing HCV core protein. Method: Heparin‐binding EGF‐like growth factor (HB‐EGF) mRNA, in HepG2 cells stably expressing HCV core protein, was detected by RT‐PCR. The soluble HB‐EGF in culture media was measured by heparin agarose chromatography/Western blot analysis. Immunodetection of Akt and IKK and IB, in HeLa cells and HepG2 cells expressing HCV core protein, were performed with neutralizing antibody for HB‐EGF, phospatidylinositol‐3‐kinase [PI(3)K] inhibitor and dominant‐negative mutant of Ras (DN‐Ras). Results: HB‐EGF expression was significantly elevated in cells expressing HCV core protein. HCV core protein activated Akt through the Ras/PI(3)K pathway by autocrine secretion of HB‐EGF. Also, HCV core protein activated IKK through Ras/PI(3)K/Akt pathway by autocrine secretion of HB‐EGF. As the Ras/PI(3)K/Akt pathway is critical in anti‐apoptotic HB‐EGF signaling, we examined the possible role of this pathway in cells expressing HCV core protein. In addition, we investigated the relationship between IB kinases (IKK) and Akt in cells expressing HCV core protein, since IKKs are known to be activated by HCV core protein and by Akt in the presence of potent mitogen. We showed that HCV core protein promoted autocrine secretion of HB‐EGF and activated Akt through the Ras/PI(3)K pathway. This model indicates a new approach to mechanism of proliferation and anti‐apoptosis in HCC. Conclusion: HCV core protein is a potent activator of mitogenic and anti‐apoptotic signaling involved in hepatocarcinogenesis.     

Clinicopathological and prognostic significance of epidermal growth factor receptor overexpression in patients with esophageal adenocarcinoma: a meta‐analysis

The prognostic significance of epidermal growth factor receptor (EGFR) overexpression in patients with esophageal adenocarcinoma (EAC) remains controversial. Eligible studies that investigated the association between survival in EAC and the expression status of EGFR were identified by an electronic search of PubMed, EMBASE, and ISI Web of Science. A meta‐analysis was performed to clarify the impact of EGFR overexpression on clinicopathological parameters or overall survival (OS) in EAC. A total of seven studies including 1028 patients were subjected to the final analysis. The overall results suggested that overexpression of EGFR was significantly correlated with not only the depth of invasion, lymph node status, and tumors stage of EAC, with a pooled odds ratio of 2.99 (95% confidence interval [CI]: 1.07–8.35; Z = 2.09; P = 0.037), 3.05 (95% CI: 1.77–5.27; Z = 4.00; P < 0.001), and 5.37 (95% CI: 2.49–11.57; Z = 4.29; P < 0.001), respectively, but also the poorer OS with a pooled hazard ratio of 2.20 (95% CI: 1.47–3.31; Z = 3.79; P < 0.001). Overexpression of EGFR correlates with not only the clinicopathological features, but also the worse OS, and it might be useful as a predictive biomarker in clinical practice, yet the clinicopathological and prognostic role of EGFR in EAC still needs further confirmation by well‐designed prospective studies.     

Insulin‐like growth factor‐I and the liver

Insulin‐like growth factors (IGFs) play an essential role in growth and development, as well as in the overall cellular regulation and metabolism in the human body. In chronic liver disease, IGF levels are decreased, and the circulating levels correlate to the extent of hepatocellular dysfunction. Patients with cirrhosis are characterised by a variety of metabolic disturbances, including nutritional and metabolic complications such as insulin resistance, malnutrition, osteopenia and hypogonadism, all related to IGF‐I deficiency. The complex process of hepatic fibrogenesis and the systemic consequences in cirrhosis are only partly understood. Disruption of the growth hormone (GH)–IGF‐I axis seems to be closely associated with the development of liver disease, and treatment with recombinant human IGF (rhIGF)‐I has been shown to halt, and even reverse, the fibrotic degeneration. IGF‐I in itself has a strong antifibrotic effect that acts directly through the GH/IGF system and indirectly by the regulation of hepatoprotective and profibrogenic factors. It is most likely that IGF‐I deficiency contributes to the diverse metabolic complications of cirrhosis. At present, liver transplantation remains the only efficient treatment of cirrhosis, and thus new methods of managing the disease are called for. RhIGF‐I supplementation and IGF‐I gene therapy may represent future perspectives of treatment.     

The circulating insulin‐like growth factor system in children with coeliac disease: an additional marker for disease activity

Background

Chronic undernutrition resulting from coeliac disease (CD) could be associated with changes in the circulating insulin‐like growth factor (IGF) system, which may participate in the pathogenesis of growth retardation occurring in these patients.

Methods

We performed a cross‐sectional study in CD subjects attempting to (1) document the pattern of serum IGF‐I and IGF binding protein (IGFBP) 1 and 3 at diagnosis and (2) assess the response of circulating IGF system to dietary treatments, in comparison with the response of clinical and laboratory findings utilized for the diagnosis of CD. Thirty‐two prepubertal CD children were divided into three groups based on the dietetic treatment: at diagnosis (D, n=18); on gluten‐free diet for at least 6 months (GFD, n=7); and on gluten challenge for at least 3 months (CH, n=7). Six postpubertal CD patients were also studied at diagnosis.

Results

In prepubertal children IGF‐I levels were significantly reduced (by 29%) in D vs sex‐ and age‐matched normal control (NC) subjects, with reductions being more pronounced before 3 years of age. Likewise, serum IGFBP‐3 concentrations were decreased by 22%, whereas circulating IGFBP‐1 levels were increased by 60%, compared with NC, with more marked IGFBP changes in older children. Similar alterations were observed in postpubertal patients. Changes in the circulating IGF system disappeared in GFD subjects and reappeared in CH children, as positivity of disease‐specific antibodies. Body mass index (BMI) also improved in GFD subjects, but did not decrease in CH children. Changes in IGF‐I and IGFBPs did not correlate with each other. Levels of IGF‐I, but not of IGFBPs, maintained the relation with age and correlated significantly with BMI and positivity of antibodies.

Conclusions

These results demonstrate that CD patients show significant changes in serum IGF‐I, in younger children, and IGFBPs (particularly IGFBP‐1), in older children and adolescents, correlating with clinical course and response to dietary treatments. The alteration in the circulating IGF system could be implicated in the pathogenesis of growth retardation occurring in CD and may provide an additional tool in monitoring of the disease. Copyright © 1999 John Wiley & Sons, Ltd.

     

FDG‐PET SUV‐max values do not correlate with epidermal growth factor receptor mutation status in lung adenocarcinoma

Previous reports suggest a correlation between positron emission tomography with fluorodeoxyglucose maximum standardized uptake value (SUVmax) and epidermal growth factor receptor (EGFR) mutation status in lung cancer. We analysed positron emission tomography with fluorodeoxyglucose SUVmax in 14 patients with EGFR mutations, and a control group of 14 subjects with wild‐type EGFR adenocarcinomas. The mean SUV value was 10.7 for EGFR‐mutated adenocarcinomas and 9.9 for wild‐type tumours. There was no correlation between SUV values and EGFR mutation status. Omitting EGFR testing in lung cancers with low SUVmax is not appropriate.