Pyoung-Han Hwang

Expression of the insulin-like growth factors (IGFs) and the IGF-binding proteins (IGFBPs) in human gastric cancer cells

Insulin-like growth factor (IGF)-I and -II are potent mitogens and postulated to exert autocrine, and paracrine effects on growth regulation in human gastric cancer. Their mitogenic effects are tightly regulated by the IGF binding proteins (IGFBPs). In this study, we evaluated the mRNA expression of IGF-I, IGF-II and the IGFBPs in a panel of human gastric cancer cell lines, and normal and tumour tissue specimens from patients with gastric cancer by reverse transcriptase-polymerase chain reaction (RT-PCR) and competitive PCR. Conditioned media (CM) of the gastric cancer cell lines were studied for the secretion of the IGFBPs by western ligand blot (WLB) and western immunoblot (WIB). IGF-I and IGF-II were expressed in all of the gastric cancer cell lines, and the normal and tumour tissue specimens. Overexpression of the IGFs, in particular, IGF-II, was observed in the tumour tissues. The expression pattern of IGFBPs was heterogeneous among the gastric cancer cell lines. IGFBP-2 was expressed in all of the gastric cancer cell lines, whereas IGFBP-1 was not detected in any cell lines. IGFBP-4 was expressed in the most of cell lines. IGFBP-3, IGFBP-5 and IGFBP-6 were expressed in approximately 50% of cell lines. In addition, exogenous IGF-I and IGF-II stimulated the proliferation of gastric cancer cells, suggesting the existence of a functional IGF system in gastric cancer. Taken together, our data-suggest that the IGF-IGFBP system may play an important role in the initiation, progression and metastasis of gastric cancer. Further studies are needed to understand the exact role of IGFs and IGFBPs in gastric neoplasia.

Anti-inflammatory effect of pachymic acid promotes odontoblastic differentiation via HO-1 in dental pulp cells

OBJECTIVES Heme oxygenase-1 (HO-1) is contributed to odontoblast differentiation in human dental pulp cells (HDPCs). In this study, pachymic acid from mushroom Formitopsis niagra is examined to determine whether it affects pulpal inflammation and promotes odontogenesis via HO-1 gene expression. MATERIALS AND METHODS The HDPCs were given H2O2 for inflammation. The anti-inflammatory character and odontoblast differentiation by pachymic acid were analyzed by Western blotting, alkaline phosphatase activity, and alizarin red S staining. To understand the mechanism of pachymic acid via HO-1 induction, the cells were treated with zinc protoporphyrin IX (ZnPP: HO-1 inhibitor). RESULTS H2O2 induced pulp inflammation and disturbed odontoblast differentiation. However, the HDPCs treated with pachymic acid affected anti-inflammatory effect and induction of odontoblast differentiation through increasing HO-1 expression. In addition, pachymic acid has potent cytoprotection and mineralization under H2O2 treatment. Furthermore, pachymic acid significantly suppressed nuclear factor-kappa B (NF-κB) translocation into nucleus and induced NE-E2-related factor-2 (Nrf2) translocation into nucleus. Overall, NF-κB and Nrf2 translocation were regulated by the HO-1 pathway. CONCLUSIONS The pachymic acid showed anti-inflammatory function and odontoblast differentiation via HO-1 pathway. These results suggested that pachymic acid may be applicable for prevention of oral inflammation or to improve dentin mineralization against several stresses.

Davallialactone from mushroom reduced premature senescence and inflammation on glucose oxidative stress in human diploid fibroblast cells.

Mushrooms are both food and a source of natural compounds of biopharmaceutical interest. The purpose of this study was to clarify whether davallialactone from mushroom extract affected the pathogenesis of hyperglycemia oxidative stress and the aging process in human diploid fibroblast (HDF) cells. The high-glucose state with glucose oxidase resulted in glucose oxidative stress, induction of inflammatory molecules, dysfunction of antioxidant molecules, and activation of mitogen-activated protein kinase (MAPKs) and its downstream signaling in old HDF cells. The exposure of glucose oxidative stress in middle-stage cells led to stress-induced premature senescence (SIPS) via senescence-associated β-galactosidase (SA β-gal) activity and displayed replicative senescence phenomena. However, davallialactone reduces the pathogenesis of glucose oxidative stress and the aging process through down-regulation of SA β-gal activity. These results strongly suggest that natural compounds, especially mushroom extract davallialactone, improve the pathogenesis of glucose oxidative stress and the aging process. Hence, davallialactone has potential in the treatment of diabetes mellitus or age-related disease complications.

Pax9 mediated cell survival in oral squamous carcinoma cell enhanced by c‐myb

Paired box gene 9 (Pax9) and c‐myb are transcription factors that regulate the expression of the genes involved in mediating cell proliferation, resistance to apoptosis, and migration. However, the function of Pax9 in oral squamous cell carcinoma (OSCC) is virtually unknown. This study examined the anti‐apoptotic roles of Pax9 and c‐myb, and clarified interaction between the two genes in KB cells. Inhibition of Pax9 caused the induction of apoptosis with enhanced cleavage of caspase‐3 and PARP, accelerated Bax, and reduced Bcl‐2 expression. Transducing c‐myb cells with adenovirus c‐myb (Ad/c‐myb) were induced cell growth and inhibited apoptosis, but dominant‐negative myb cells (Ad/DN‐myb) were not affected. Pax9 was upregulated in the Ad/c‐myb cells with simultaneous decrease in the Ad/DN‐myb infection. However, c‐myb remained unaffected in the Pax9 small interfering RNA (siRNA) transfected cells. Moreover, the Pax9 siRNA transfected cells and Ad/DN‐myb infected cells were able to arrest the cell cycle at the G0 phase. This suggests that Pax9 and c‐myb expression in KB cells is essential for cell growth, and survival is enhanced by c‐myb. Disrupting the function of c‐myb and Pax9 could be a potential target for cancer treatment. Copyright

Amphiphilic triblock copolymer poly(p-dioxanone-co-L-lactide)-block-poly(ethylene glycol), enhancement of gene expression and inhibition of lung metastasis by aerosol delivery.

We describe the development of an aerosol system for topical gene delivery to the lungs of C57BL/6 mice. This system is based on the combination of the commercial cationic lipid Lipofectin with a novel amphiphilic triblock copolymer, poly(p-dioxanone-co-L-lactide)-block-poly(ethylene glycol) (PPDO/PLLA-b-PEG, and abbreviated in the text as polymeric micelles). After optimizing conditions for DNA delivery to the lungs of mice using the combination of polymeric micelles with Lipofectin and LacZ DNA, we used the Lipofectin/polymeric micelle system to deliver the tumor suppressor gene PTEN to the lungs of C57BL/6 mice bearing the B16-F10 melanoma. Lipofectin/PTEN/polymeric micelles significantly improved gene expression of PTEN in the lungs of mice with no evidence of cell toxicity or acute inflammation. Importantly, lung metastasis, as measured by lung weight, was significantly reduced (P<0.001), as were total tumor foci in the lungs (P<0.001) and size of individual tumor nodules in animals treated with Lipofectin/PTEN/polymeric micelles compared with control animals. Survival time was also extended. These results suggest that the Lipofectin/polymeric micelle system is appropriate for enhancing gene delivery in vivo and that it can be applied as a non-invasive gene therapy for lung cancer.

Protein Kinase CK2/PTEN Pathway Plays a Key Role in Platelet-Activating Factor-Mediated Murine Anaphylactic Shock

Platelet-activating factor (PAF) is a major mediator in the induction of fatal hypovolemic shock in murine anaphylaxis. This PAF-mediated effect has been reported to be associated with PI3K/Akt-dependent eNOS-derived NO. The phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is phosphatidylinositol phosphate phosphatase, which negatively controls PI3K by dephosphorylating the signaling lipid, phosphatidylinositol 3,4,5-triphosphate. In this study, we examined the possible involvement of PTEN in PAF-mediated anaphylactic shock. Induction of anaphylaxis or PAF injection resulted in a rapid decrease in PTEN activity, followed by increases in PI3K activity and phosphorylation of Akt and eNOS. Systemic administration of adenoviruses carrying PTEN cDNA (adenoviral PTEN), but not the control AdLacZ, not only attenuated anaphylactic symptoms, but also reversed anaphylaxis- or PAF-induced changes in PTEN and PI3K activities, as well as phosphorylation of Akt and eNOS. We found that the decreased PTEN activity was associated with PTEN phosphorylation, the latter effect being prevented by the protein kinase CK2 inhibitor, DMAT. DMAT also inhibited anaphylactic symptoms as well as the anaphylaxis- or PAF-mediated PTEN/PI3K/Akt/eNOS signaling cascade. CK2 activity was increased by PAF. The present data provide, as the key mechanism underlying anaphylactic shock, PAF triggers the upstream pathway CK2/PTEN, which ultimately leads to the activation of PI3K/Akt/eNOS. Therefore, CK2/PTEN may be a potent target in the control of anaphylaxis and other many PAF-mediated pathologic conditions.

c-myb stimulates cell growth by regulation of insulin-like growth factor (IGF) and IGF-binding protein-3 in K562 leukemia cells

c-myb plays an important role in the regulation of cell growth and differentiation, and is highly expressed in immature hematopoietic cells. The human chronic myelogenous leukemia cell K562, highly expresses IGF-I, IGF-II, IGF-IR, and IGF-induced cellular proliferation is mediated by IGF-IR. To characterize the impact of c-myb on the IGF-IGFBP-3 axis in leukemia cells, we overexpressed c-myb using an adenovirus gene transfer system in K562 cells. The overexpression of c-myb induced cell proliferation, compared to control, and c-myb induced cell growth was inhibited by anti-IGF-IR antibodies. c-myb overexpression resulted in a significant increase in the expression of IGF-I, IGF-II, and IGF-IR, and a decrease in IGFBP-3 expression. By contrast, disruption of c-myb function by DN-myb overexpression resulted in significant reduction of IGF-I, IGF-II, IGF-IR, and elevation of IGFBP-3 expression. In addition, exogenous IGFBP-3 inhibited the proliferation of K562 cells, and c-myb induced cell growth was blocked by IGFBP-3 overexpression in a dose-dependent manner. The growth-promoting effects of c-myb were mediated through two major intracellular signaling pathways, Akt and Erk. Activation of Akt and Erk by c-myb was completely blocked by IGF-IR and IGFBP-3 antibodies. These findings suggest that c-myb stimulates cell growth, in part, by regulating expression of the components of IGF-IGFBP axis in K562 cells. In addition, disruption of c-myb function by DN-myb may provide a useful strategy for treatment of leukemia.

c‐myb mediates inflammatory reaction against oxidative stress in human breast cancer cell line, MCF‐7

Emerging evidence suggests that oncogenes play an important role in the inflammatory reactions in cancer cells, but the precise molecular and cellular mechanisms linking the oncogenes to inflammation is unclear. This study examined the contribution of proto‐oncogene c‐myb to inflammation in MCF‐7 breast cancer cells.

Adriamycin inhibits adipogenesis through the modulation of PPARγ and restoration of adriamycin-mediated inhibition of adipogenesis by PPARγ over-expression

Adriamycin is an anti-cancer drug, effective against a wide range of cancers. However, its clinical application is limited by its cardiotoxicity. A number of reports suggest that adriamycin induces bodyweight loss also. The aim of this study was to investigate the effect of adriamycin on adipogenesis as bodyweight chancges can be directly correlated with adipocytes. Fat accumulation in 3T3-L1 pre-adipocytes, as a result of adipogenesis was detected using oil red O staining. We performed western immunoblot for the expression of adipocyte differentiation related genes to analyze the molecular mechanism of adriamycin-mediated inhibition of adipogenesis. Over-expression of target gene was done by using recombinant adenoviruses. Adriamycin inhibited adipogenesis in a dose-dependent manner. It was observed that adriamycin down-regulated the expression of PPARγ. Moreover, up-stream elements of PPARγ were also found to be down-regulated by adriamycin. Adriamycin might prevent bodyweight gain through inbibition of adipogenesis by the down-regulation of PPARγ and its up-stream transcriptional regulators like C/EBPβ and KLF4. To reverse the adriamycin-mediated inhibition of adipogenesis, PPARγ was over-expressed by adenoviral mediated gene delivery. Over-expression of PPARγ partially restored adipogenesis. Moreover, the early regulators of adipogenesis were also found to be restored after the over-expression of PPARγ. Adriamycin down-regulates the expression of PPARγ which leads to prevention of bodyweight gain through inhibition of adipogenesis. Activation of PPARγ by either adenoviral mediated gene delivery or by using PPARγ agonist may be useful in controlling the bodyweight loss.

Clinical and hematologic manifestations in patients with Diamond Blackfan anemia in Korea.

Background Diamond Blackfan anemia (DBA), characterized by impaired red cell production, is a rare condition that is usually symptomatic in early infancy. The purpose of this study was to assess nationwide experiences of DBA encountered over a period of 20 years. Methods The medical records of 56 patients diagnosed with DBA were retrospectively reviewed from November 1984 to July 2010. Fifteen institutions, including 13 university hospitals, participated in this study. Results The male-to-female ratio of patients with DBA was 1.67:1. The median age of diagnosis was 4 months, and 74.1% were diagnosed before 1 year of age. From 2000 to 2009, annual incidence was 6.6 cases per million. Excluding growth retardation, 38.2% showed congenital defects: thumb deformities, ptosis, coarctation of aorta, ventricular septal defect, strabismus, etc. The mean hemoglobin concentration was 5.1±1.9 g/dL, mean corpuscular volume was 93.4±11.6 fL, and mean number of reticulocytes was 19,700/mm3. The mean cellularity of bone marrow was 75%, with myeloid:erythroid ratio of 20.4:1. After remission, 48.9% of patients did not need further steroids. Five patients with DBA who received hematopoietic transplantation have survived. Cancer developed in 2 cases (3.6%). Conclusion The incidence of DBA is similar to data already published, but our study had a male predilection. Although all patients responded to initial treatment with steroids, about half needed further steroids after remission. It is necessary to collect further data, including information regarding management pathways, from nationwide DBA registries, along with data on molecular analyses.