The Tumor Suppressor Function of PTEN/MMAC1 through the Regulation of IGFs and IGFBPs / 소아과

PURPOSE: PTEN/MMAC1, a novel tumor suppressor gene, is mutated in a variety of advanced and metastatic cancers. It acts as a phosphatase, and thereby, regulates the PI-3 kinase/Akt pathway. In this study, we examined to evaluate the new function of anti-tumor effects of PTEN/MMAC1 through the regulation of the IGFs-IGFBPs in gastric cancer cells. METHODS: PTEN/MMAC1 was expressed in an adenovirus-mediated gene delivery system and introduced into gastric cancer cells(SNU-484 & SNU-668) in vitro. The effect of cell growth and the expression of IGFs and IGFBPs after Ad/PTEN infection was analyzed by MTT assay, RT-PCR and Western immunoblot. RESULTS: Ad/PTEN infected cells were inhibited in cell growth compared with moak cells and Ad/ LacZ infected cells. Overexpression of PTEN/MMAC1 induced decrease in expression of IGF-I, -II and IGF-I receptors which are known as growth prompt molecules in a variety of cancers. Of the six IGFBPs, the expressions of IGFBP-4 and IGFBP-6 were decreased in Ad/PTEN infected cells. In contrast, IGFBP-3 expression was markedly increased by up to 3-fold in Ad/PTEN infected cells. Overexpression of PTEN/MMAC1 inhibited the activation of Akt/PKB pathway, but had no effect on the MAPK pathway. CONCLUSION: These findings suggest that the tumor suppressor function of PTEN/MMAC1 is, at least in part, mediated through the down-regulation of IGF-I abd IGF-II, and up-regulation of IGFBP-3 in gastric cancer cells by the inhibition of PI-3 kinase pathway.

Diagnostic Approach of Wiskott-Aldrich Syndrome / 소아과

OBJECTIVE: A probable diagnosis of Wiskott-Aldrich syndrome(WAS) should be considered in any boy presenting with unusual bleeding, congenital thrombocytopenia and small platelets. The definitive diagnosis of WAS is usually made by the detection of WASP gene mutation or a decrease or absence of the WAS protein(WASP) in blood cells using molecular genetic analysis. However, these methods are too time-consuming and difficult. In this study, therefore, we tried to compare various diagnostic methods and establish the molecular screening steps for the definitive diagnosis of WAS. METHODS: Peripheral blood was drawn from WAS patient with clinical characteristic symptoms, and analyzed with automated complete blood cell count analysis, immunological analysis, and molecular analysis. The morphologic change of WAS patient's blood cell membrane was examined by scanning electron microscopy(SEM). Protein analysis for the expression of WASP protein in PBMC cells was evaluated by FACS and Western immunoblot. Genetic analysis for the detection of a mutation of the WASP gene was performed by polymerase chain reaction-single strand conformational polymorphism analysis(PCR-SSCP) and direct sequencing of PCR products. RESULTS: In addition to microthrombocytopenia, our investigation revealed morphologic defects in WAS lymphocytes by SEM and abnormal mobility shifting in WAS patients by PCR-SSCP. Sequencing the WASP gene detected a specific single base mutation in exon 2, resulting in missense substitution of adenine for guanine 208(G208A, Gly70Arg). FACS and Western immunoblot demonstrated absent expressions of WASP in WAS patients and reduced expression of WASP in carrier when compared with normal controls. CONCLUSION: From these results, we suggested the following diagnostic approaches in patients suspected of having WAS. Protein-based analysis such as FACS and Western immunoblot should be employed as the first line of investigation. The second line genetic analyses should employ second- step approaches with localization of mutation by screening exons, typically by PCR-SSCP, followed by direct sequencing.

Serum Ghrelin Concentrations in Type 2 Diabetes Mellitus / 대한소아내분비학회지

PURPOSE:Ghrelin is an endogenous ligand of the growth hormone secretagogue receptor, and stomach is the major site of ghrelin secretion. The purpose of this study is to compare the serum ghrelin concentrations between patients with type 2 diabetes mellitus (DM) and normal adults. We studied also whether serum ghrelin levels in the patients with type 2 DM are correlated with body mass index (BMI), serum insulin, lipid profiles, and creatinine levels. METHODS:Forty patients with type 2 DM and forty normal adults were included in this study. We measured heights and weights of the subjects and calculated their BMIs. Blood samples were obtained to measure the ghrelin concentration and their sera were stored at -20degreeC until used. In all subjects, serum ghrelin levels were measured using the commercially available Ghrelin(human) EIA kit. RESULTS:No differences of mean values were detected between the control group and the type 2 diabetic group for age, body weight, BMI, and the levels of serum total cholesterol, triglyceride, HDL cholesterol, and creatinine. But ghrelin level of the type 2 diabetic group (71.1+/-30.5 ng/L) was significantly lower than the control group (139.7+/-36.9 ng/L). In the control group, the ghrelin level showed positive correlation with HDL cholesterol (Pearson's correlation coefficient=0.37, P<0.05). In the diabetic group, the ghrelin level showed weakly positive correlation with insulin concentration. However, there was no significant relationship between serum ghrelin and various parameters in the diabetic patients group. CONCLUSION: In this study, ghrelin concentration in type 2 diabetic patients was lower than that in the control group. In the control group, serum ghrelin concentrations were positively correlated with HDL cholesterol. In the type 2 diabetic group, there was no significant correlation between insulin and ghrelin concentrations.

Novel Gap Junction Molecules, Connexin 37, Enhances the Bystander Effect in HSVtk/GCV Gene Therapy

PURPOSE: Gap junction intercellular communication(GJIC) is an important mechanism of the bystander effect in herpes simplex thymidine kinase/ganciclovir(HSVtk/GCV) gene therapy Therefore, we attempted to enhance the bystander effect in vitro by exogenous overexpressing connexin 37(Cx37) in cells to increase GJIC. METHODS: NIH3T3 cells were transfected with the Cx37 and HSVtk gene or the HSVtk gene alone by the calcium phosphate method, and we detected their expression from these cells by RT-PCR. GCV-mediated cytotoxicity and the bystander effect of each transfectant was then assessed and compared. RESULTS: Cells transfected with HSVtk became sensitive to low concentration of GCV. We found significantly increased cytotoxicity in HSVtk/GCV gene therapy after introduction of the HSVtk and Cx37 genes together compared with the cytotoxicity seen after introduction of the HSVtk gene in vitro. Co-expression of the HSVtk and Cx37 genes potentiates HSVtk/GCV gene therapy through the bystander effect. CONCLUSION: These results indicated that the increase of GJIC using Cx37 have potentiated the by stander effect of HSVtk/GCV therapy, and may be a new approach to improve response in suicidal cancer gene therapy.

A Case of Cerebral Vasculitis in Henoch-Sh nlein Purpura

Henoch-Sh nlein Purpura(HSP) is an immuologically mediated systemic vasculitis of small blood vessels affecting predominantly the skin, gastrointestinal tract, joints, and kidneys. Clinical neurological manifestations such as headaches, behavioral changes, mental changes, seizures, and visual loss are described, but neurological complication are rare during the course of HSP. We experienced a case of an 8 year-old male with HSP who presented with seizures. Magnetic resonance imaging(MRI) showed multiple high signal intensity in both cortical and subcortical areas of frontal and parieto-occipital lobes and magnetic resonance(MR) angiogram showed stenosis of cerebral arteries, compatible with MRI and MR angiogram findings of cerebral vasculitis. We report this case with related literature.