[Show abstract][Hide abstract] ABSTRACT: Angiogenesis is considered essential for proper bone regeneration. The purpose of this investigation was to determine if a combined therapy of bone morphogenetic protein-2 (BMP-2) and cartilage oligomeric matrix protein angiopoietin-1 (COMP-Ang1) can potentiate the therapeutic effect of BMP-2 in a rat model of ischemic necrosis of the femoral head (INFH). INFH was surgically induced in the femoral head of rats, and the animals were divided into the following groups: 1) a sham-operated group (sham group), 2) a bovine serum albumin-injected group (BSA group), 3) a BMP-2-injected group (BMP-2 group), and 4) a COMP-Ang1 and BMP-2-injected group (COMP-Ang1 + BMP-2 group) (n = 20/group). Radiologic, histologic, and histomorphometric assessments were performed to assess femoral head morphology, vascular density, and bone resorption activity. Western blots and immunohistochemical staining were performed to evaluate production of BMP-related signaling proteins in C3H10T1/2 cells and tissues. Real-time RT-PCR was performed to investigate expression of the target integrin gene, and the effect of integrin on C3H10T1/2 cells was determined using a cell adhesion assay. Radiographs obtained six weeks after injection revealed better preservation of the architecture of the femoral head in the COMP-Ang1 + BMP-2 group compared with the BSA and BMP-2 groups. Histological findings indicated increased trabecular bone and vascularity and decreased osteoclast bone resorption activity in the COMP-Ang1 + BMP-2 group compared with those in the BSA and BMP-2 groups. The combination of COMP-Ang1 and BMP-2 increased phosphorylation of Smad1/3/5, p38, and Akt. Increased integrin α3 and β1 mRNA expression in the COMP-Ang1 + BMP-2 group promoted cell adhesion. These results suggest that COMP-Ang1 preserved the necrotic femoral head through the potentiation of BMP-2 signaling pathways and angiogenesis. Combination treatment with COMP-Ang1 and BMP-2 may be a clinically useful therapeutic application in INFH.
PLoS ONE 10/2014; 9(10):e110593. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: SIRT1 is a class III histone deacetylase that plays diverse roles in various cancers. However, the clinical significance of SIRT1 in hepatocellular carcinoma (HCC) remains unknown.
Journal of Hepatology 08/2014; · 10.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Type 1 diabetes is an autoimmune disease caused by permanent destruction of insulin-producing pancreatic β cells and requires lifelong exogenous insulin therapy. Recently, islet transplantation has been developed, and although there have been significant advances, this approach is not widely used clinically due to the poor survival rate of the engrafted islets. We hypothesized that improving survival of engrafted islets through ex vivo genetic engineering could be a novel strategy for successful islet transplantation. We transduced islets with adenoviruses expressing betacellulin, an epidermal growth factor receptor ligand, which promotes β-cell growth and differentiation, and transplanted these islets under the renal capsule of streptozotocin-induced diabetic mice. Transplantation with betacellulin-transduced islets resulted in prolonged normoglycemia and improved glucose tolerance compared with those of control virus-transduced islets. In addition, increased microvascular density was evident in the implanted islets, concomitant with increased endothelial von Willebrand factor immunoreactivity. Finally, cultured islets transduced with betacellulin displayed increased proliferation, reduced apoptosis and enhanced glucose-stimulated insulin secretion in the presence of cytokines. These experiments suggest that transplantation with betacellulin-transduced islets extends islet survival and preserves functional islet mass, leading to a therapeutic benefit in type 1 diabetes.
Experimental and Molecular Medicine 05/2014; 46:e98. · 2.46 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: After intramembranous proteolysis-mediated loss of the extracellular domain of the epithelial cell adhesion molecule (EpEx) and release of an intracellular domain (EpICD) into the cytoplasm, EpICD sequentially associates with FHL2 to form a nuclear complex with β-catenin and Lef-1. This association induces gene transcription involved in the activation of the oncogenic potential of epithelial cell adhesion molecule (EpCAM). We examined the localization and expression of EpEx, EpICD and β-catenin in surgical specimens of extrahepatic cholangiocarcinoma (ECC) from 79 patients and focused on the relationship between nuclear expression of EpICD and β-catenin. We also examined the role of EpICD by transfecting the EpICD cDNA in cholangiocarcinoma (CC) cell lines. There was a significant correlation between the nuclear expression of EpICD and β-catenin in ECC tissues. Frequent nuclear co-localization of EpICD and β-catenin was observed in cancer cells forming the invasive front. Nuclear expression of EpICD also significantly correlated with histologic grade of tumor. Overexpression of EpICD in the CC cells significantly increased the cell growth and proliferation. The overexpression of EpICD in the CC cells also increased the expression levels of the active form of β-catenin and EpCAM target genes, such as c-myc and cyclin D1. Furthermore, the overexpression of EpICD significantly enhanced the migration and invasiveness of CC cells. Conversely, the inhibition of EpCAM in EpCAM-overexpressing cells by siRNA significantly decreased cell proliferation, migration and invasion. These results indicate that the spatial localization of EpICD and its mutual interaction with β-catenin may be important in ECC progression and invasion.
International Journal of Oncology 05/2014; · 2.77 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Insulin-like growth factor binding protein 3 (IGFBP-3) is known to interfere with the NF-κB signaling pathway, and it effectively promotes apoptosis in tumor cells by a variety of mechanisms. NF-κB activation and apoptosis resistance of fibroblast-like synoviocytes (FLS) play pivotal roles in rheumatoid arthritis (RA). This study was undertaken to evaluate whether IGFBP-3 has antiarthritic effects.
To deliver IGFBP-3, we used an adenovirus containing IGFBP-3 complementary DNA (AdIGFBP-3) or IGFBP-3 mutant that is devoid of IGF binding affinity but retains IGFBP-3 receptor binding ability (AdmtIGFBP-3). The regulatory roles of IGFBP-3 in inflammation and bone destruction were investigated in mice with collagen-induced arthritis (CIA).
IGFBP-3 levels were significantly higher in patients with RA than in those with osteoarthritis (OA) and were notably higher in patients with active RA. AdIGFBP-3 suppressed NF-κB activation, chemokine production, and matrix metalloproteinase secretion induced by tumor necrosis factor α (TNFα) in RA FLS. AdIGFBP-3 sensitized RA FLS to TNFα-induced apoptosis in vitro and also significantly increased apoptosis in an in vivo model of Matrigel implants engrafted into immunodeficient mice. AdIGFBP-3-injected mice with CIA had attenuated arthritis severity and reduced radiologic and pathologic abnormalities. Moreover, AdIGFBP-3 down-regulated local and systemic levels of NF-κB-targeted proinflammatory cytokines. Of note, RA FLS and mice with CIA treated with AdmtIGFBP-3 exhibited similar effects as those treated with AdIGFBP-3.
Our results suggest that both the inflammatory response and bone destruction are reduced with blockage of NF-κB activation and induction of apoptosis in RA FLS by IGFBP-3. Therefore, IGFBP-3 may have therapeutic potential in RA.
[Show abstract][Hide abstract] ABSTRACT: Platelet-activating factor (PAF) promotes tumor metastasis via activation of the transcription factor NF-κB. We here investigated the role of the protein kinase CK2 in PAF-induced NF-κB activation and tumor metastasis, given that PAF has been reported to increase CK2 activity, and that CK2 plays a key role in NF-κB activation. PAF increased CK2 activity, phosphorylation and protein expression in vivo as well as in vitro. CK2 inhibitors inhibited the PAF-mediated NF-κB activation and expression of NF-κB-dependent proinflammatory cytokines and antiapoptotic factors. Pretreatment with the antioxidant NAC resulted in a significant inhibition in PAF-induced enhancement of CK2 activity, phosphorylation, and protein expression in vivo as well as in vitro. H2O2 and known reactive oxygen species (ROS) inducers, LPS and TNF-α enhanced CK2 activity, phosphorylation, and protein expression, which was again inhibited by antioxidant. PAF-, LPS- and TNF-α-induced increased CK2 activity, phosphorylation, and protein expression, which were inhibited by p38 inhibitor. PAF, LPS or TNF-α increased pulmonary metastasis of B16F10, which was inhibited by antioxidants, CK2 inhibitor and p38 inhibitor. Our data suggest that 1) ROS activates CK2 via p38, which, in turn, induces NF-κB activation, and 2) PAF, LPS and TNF-α increase pulmonary tumor metastasis via the induction of the ROS/p38/CK2/NF-κB pathway.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Musashi-1 (Msi-1) is proposed to be a marker of progenitor cells in the human gastric mucosa. We examined Msi-1 expression and localization in surgical specimens of gastric cancer in young patients using immunohistochemistry and tested associations of Msi-1 expression with clinicopathological features. Patients (n=611) with gastric cancer who underwent radical gastrectomy were included in the present study. To minimize confounding effects, we matched 26 gastric cancer patients 30 years of age or younger (age ≤30) with 26 patients 60 years of age or older (age ≥60). The groups were matched by gender, tumor histological type and tumor stage. Gastric cancer in the younger patients was significantly associated with female gender and with diffuse histological type, compared with 585 gastric cancer patients older than 31 years. Msi-1 expression was more frequently upregulated in gastric cancer in young patients than in patients older than 60 years. Msi-1 expression was significantly associated with diffuse histological type, depth of tumor invasion, lymph node metastasis and tumor stage in the 26 young patients with gastric cancer. Univariate Kaplan‑Meier survival analysis identified Msi-1 expression as a significantly negative factor in the survival of young gastric cancer patients. However, Msi-1 expression was not significantly associated with survival in the 26 matched older patients. According to mucin phenotype, the gastric foveolar type predominated in Msi-1-positive gastric cancers. Our principal findings included a significantly higher level of Msi-1 expression in younger gastric cancer patients compared to older ones, and a probable association of tumor Msi-1 expression in young gastric cancer patients with more aggressive tumor type.
International Journal of Oncology 01/2014; · 2.77 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The phosphorylation of proteins on serine/threonine residues that immediately precede proline (pSer/Thr-Pro) is a key signaling mechanism by which cell cycle regulation and cell differentiation and proliferation occur. The peptidyl-prolyl isomerase PIN1-catalyzed conformational changes of the pSer/Thr-Pro motifs may have profound effects on the function of numerous oncogenic and cell signaling pathways. To date, no studies have examined the expression of PIN1 and its potential role in the pathogenesis of extrahepatic cholangiocarcinoma (ECC). Therefore, the present study performed an immunohistochemistry analysis of the expression of PIN1 in 67 cases of ECC and evaluated its association with clinicopathological factors. In addition, the role of PIN1 was examined using synthetic small interfering RNA (siRNA) to silence PIN1 gene expression in human CC RBE cells. Positive PIN1 expression was observed in 35 of the 67 (52.2%) ECC cases and was predominantly localized to the nucleus of the tumor cells. The immunoreactive score for PIN1 was significantly higher in the tumor cells (4.07±0.4) compared with the adjacent benign bile duct cells (1.19±0.4) (P<0.001). PIN1 expression was significantly correlated with tumor cell proliferation (Ki-67 labeling index; P=0.024). Silencing PIN1 expression using siRNA significantly decreased the proliferation, migration and invasion of the tumor cells. In conclusion, the results indicated that the expression of PIN1 may play a key role in the development and progression of ECC.
[Show abstract][Hide abstract] ABSTRACT: Nerve growth factor (NGF) is a neurotrophin and has been suggested to induce heme oxygenase-1 (HO1) expression. Although the role of HO1 in tumorigenesis remains controversial, recent evidence suggests NGF and HO1 as tumor-progressing factors. However, the correlative role of NGF and HO1 and their prognostic impact in breast carcinoma is unknown.
We investigated the expression and prognostic significance of the expression of NGF and HO1 in 145 cases of breast carcinoma.
Immunohistochemical expression of NGF and HO1 was observed in 31% and 49% of breast carcinoma, respectively. The expression of NGF and HO1 significantly associated with each other, and both have a significant association with histologic grade, HER2 expression, and latent distant metastasis. The expression of NGF and HO1 predicted shorter overall survival of breast carcinoma by univariate and multivariate analysis. NGF expression was an independent prognostic indicator for relapse-free survival by multivariate analysis. The combined expression pattern of NGF and HO1 was also an independent prognostic indicator of overall survival and relapse-free survival. The patients with tumors expressing NGF had the shortest survival and the patients with tumor, which did not express NGF or HO1 showed the longest survival time.
This study has demonstrated that individual expression of NGF or HO1, and the combined NGF/HO1 expression pattern could be prognostic indicators for breast carcinoma patients.
BMC Cancer 11/2013; 13(1):516. · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The epithelial to mesenchymal transition (EMT) is a crucial process in tumor progression. EMT of tumor cells not only causes increased metastasis, but also contributes to drug resistance. Serum response factor (SRF) is a transcription factor that plays a central role in carcinogenesis and tumor progression in several types of cancers. We investigated the effect of EMT-related SRF, focusing on its promotion of chemoresistance against sorafenib in hepatocellular carcinoma (HCC). We examined SRF and Snail expression in 146 cases of HCCs by immunohistochemistry. We also examined the chemoresistance effect of SRF in HCC cells by transfecting HLE cells with SRF cDNA and SH-J1 cells with SRF antisense cDNA. Expression of SRF and Snail were detected in 37.6% (55 of 146 cases) and in 12.3% (18 of 146 cases) of the HCCs, respectively. None of the tumor-free liver tissues showed SRF or Snail expression. SRF expression was closely correlated with the expression of Snail (p<0.001) and expression of both SRF and Snail showed significant correlation with the high histological grade (p=0.015 and 0.003, respectively). Overexpression of SRF in HLE cells led to increased expression of mesenchymal markers, as well as increased cell growth and colony formation. Overexpression of SRF also led to a significant reduction in the cytotoxic effect of sorafenib in HLE cells. Conversely, inhibition of SRF expression in the SH-J1 cells significantly enhanced the apoptotic effects of sorafenib, along with the reduced expression of mesenchymal markers and restored the expression of E-cadherin. These results suggest that SRF is critical for HCC to acquire a mesenchymal phenotype, which leads to resistance against a sorafenib-mediated apoptotic effect.
International Journal of Oncology 10/2013; · 2.77 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recently, the important role of silent mating type information regulation 2 homolog 1 (SIRT1) and deleted in breast cancer 1 (DBC1) in human cancer has been extensively studied and their role has been closely related with the control of P53 and androgen receptor (AR) functions. However, their role in clear cell renal cell carcinoma (CRCC) is still unknown.
We evaluated the expression of SIRT1, P53, acetylated-P53, DBC1 and AR and their prognostic significance in 200 CRCC patients.
The expression of SIRT1, P53, DBC1, and AR significantly correlated with each other and all of them predicted shorter overall survival (OS), relapse-free survival (RFS), and cancer-specific survival (CSS). In contrast, the expression of acetylated-P53 predicted favourable OS, RFS, and CSS. Combined expression pattern of acetylated-P53 and P53 (Ac-P53/P53) also closely correlated with survival of CRCC patients. Multivariate analysis revealed DBC1, acetylated-P53, and Ac-P53/P53 expression as independent prognostic indicators for OS and RFS, and Ac-P53 expression as an independent prognostic indicator for CSS.
This study demonstrates that the acetylation status of P53 and the expression of SIRT1, DBC1, and AR could be new prognostic indicators for CRCC and suggest that SIRT1-P53 and DBC1-AR related pathways could be new therapeutic targets for the treatment of CRCC.
[Show abstract][Hide abstract] ABSTRACT: We investigated the clinical significance of epithelial-mesenchymal transition (EMT) phenotype in 184 small intestinal adenocarcinomas (SIACs) based on the expression pattern of EMT-related proteins in cancer cells.
Immunohistochemistry for epithelial (E-cadherin) and mesenchymal (vimentin and fibronectin) markers were performed and cases of SIAC were classified into four subtypes of EMT: complete type (E-cadherin-, vimentin+ and/or fibronectin+), wild type (E-cadherin+, vimentin-, fibronectin-), incomplete 1 type (hybrid type; E-cadherin+, vimentin+ and/or fibronectin+), and incomplete 2 type (null type; E-cadherin-, vimentin-, fibronectin-).
We identified 19 (10.3%) cases of complete EMT type, 86 (46.7%) cases of wild type and 79 (43%) cases of incomplete EMT type [hybrid type, 22 (12%) cases; null type, 57 (31%) cases]. Complete EMT phenotype showed a significant association with undifferentiated histology (p < 0.001). Overall survival of SIAC patients with complete EMT phenotype was significantly shorter than those of patients with incomplete (p = 0.001) and wild (p < 0.001) types. In multivariate analysis, complete EMT phenotype was an independent prognostic factor in SIAC patients (hazard ratio 2.3; 95% confidence interval 1.15-4.6; p = 0.019).
Complete EMT phenotype stratifies a specific group representing a poor clinical outcome in patients with SIAC.
[Show abstract][Hide abstract] ABSTRACT: Recently, the roles of SIRT1 and deleted in breast cancer 1 (DBC1) in human cancer have been extensively studied and it has been demonstrated that they are involved in many human carcinomas. However, their clinical significance for soft-tissue sarcomas has not been examined. In this study, we evaluated the expression and prognostic significance of the expression of SIRT1, DBC1, P53, β-catenin, cyclin D1, and KI67 in 104 cases of soft-tissue sarcomas.
Immunohistochemical expression of SIRT1, DBC1, P53, β-catenin, and cyclin D1 were seen in 71%, 74%, 53%, 48%, and 73% of sarcomas, respectively. The expression of SIRT1, DBC1, P53, β-catenin, and cyclin D1 were significantly correlated with advanced clinicopathological parameters such as higher clinical stage, higher histological grade, increased mitotic counts, and distant metastasis. The expression of SIRT1, DBC1, P53, β-catenin, cyclin D1, and KI67 were significantly correlated with each other and positive expression of all of these predicted shorter overall survival and event-free survival by univariate analysis. Multivariate analysis revealed the expression of SIRT1 as an independent prognostic indicator for overall survival and event-free survival of sarcoma patients. In conclusion, this study demonstrates that SIRT1- and DBC1-related pathways may be involved in the progression of soft-tissue sarcomas and can be used as clinically significant prognostic indicators for sarcoma patients. Moreover, the SIRT1- and DBC1-related pathways could be new therapeutic targets for the treatment of sarcomas.
PLoS ONE 09/2013; 8(9):e74738. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Angiogenesis plays a critical role in synovial inflammation and joint destruction in rheumatoid arthritis (RA). The vascular endothelial growth factor (VEGF)-A and angiopoietins are two important mediators of synovial angiogenesis. We have previously developed a novel chimeric decoy receptor, namely double anti-angiogenic protein (DAAP), which can bind both VEGF-A and angiopoietins, blocking their actions. This study was performed to evaluate the anti-arthritic effect of DAAP and combination effect with the TNF- inhibitor in collagen-induced arthritis (CIA).
Recombinant DAAP, VEGF-Trap, Tie2-Fc, and dimeric-Fc proteins were produced and purified from CHO cells in large-scale bioreactors. CIA was induced in DBA/1 mice with type II collagen. The preventive effect of DAAP was determined and compared with other decoy receptors such as VEGF-Trap or Tie2-Fc, which block VEGF-A or angiopoietins, respectively. The clinical, radiographic, pathologic, and immunohistochemical analyses were performed in CIA mice. The levels of matrix metalloprotease-3 (MMP-3) and interleukin-1 (IL-1) were quantified by ELISA, and receptor activator of nuclear factor-kappa B ligand (RANKL) mRNA levels were measured with polymerase chain reaction (PCR). Finally, we investigated the combination effects of DAAP with low dose of TNF-alpha decoy receptor (etanercept, 10 mg/kg).
DAAP had a much greater inhibitory effect on arthritis severity and bone destruction than VEGF-Trap or Tie2-Fc on clinical and radiographic evaluation. These inhibitory effects were accompanied by significantly diminishing pathologic abnormalities, CD31-positive vasculature, and synovial infiltration by F4/80-positive macrophages. The levels of MMP-3, IL-1, and RANKL were much lower in DAAP-injected group than those of control. Furthermore, DAAP showed therapeutic effect and a combination effect with etanercept when injected after arthritis onset in established CIA.
DAAP has not only potent prophylactic effects on both inflammation and bone destruction, but also therapeutic effects, alone and combination effects with a TNF- inhibitor in CIA mice. These results suggest that DAAP could be used as an effective new therapeutic agent for RA.
Arthritis research & therapy 08/2013; 15(4):R85. · 4.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The pseudoreceptor BAMBI (bone morphogenetic protein and activin membrane-bound inhibitor), formerly known as NMA, is an inhibitor of the TGF-β signaling pathway. BAMBI exhibits structural homology to TGF-βRI but lacks an intracellular kinase domain. In most of the high-grade carcinomas, the degree of BAMBI expression is abnormally increased, which leads to the proliferation and metastasis of tumor cells. Recent studies have reported that BAMBI is involved in the Wnt-β-catenin pathway that regulates the proliferation and metastasis of tumor cells. However, little is known about the role of BAMBI and β-catenin in human osteosarcoma. Given the above background, we examined the role of BAMBI in the pathophysiology of osteosarcoma. Using immunohistochemical staining and western blot analysis, the degree of the expression of BAMBI and β-catenin was significantly higher in osteosarcoma specimens compared with normal tissues. With the overexpression of BAMBI, mediated by adenovirus, the degree of invasion and migration was significantly increased and the proliferation of U2-OS osteosarcoma cells was stimulated. Transwell analysis showed that BAMBI increased the invasion of osteosarcoma cells and upregulated the secretion of matrix metalloproteinases (MMPs), which was demonstrated by gelatin zymography. Fluorescence-activated cell sorting (FACS) analysis showed a significant arrest in cell cycle progression at G0/G1 in osteosarcoma cells transfected with siRNA targeting BAMBI. With the overexpression of BAMBI, mediated by the adenovirus, however, there was a decrease in the number of cells at G0/G1. Consistent with the findings that cell growth was increased, BAMBI promoted the transition from G0/G1 to G2/M in the osteosarcoma cells. Our results suggest that BAMBI plays a key role in the pathogenesis and progression of osteosarcoma by regulating the expression of β-catenin and other signaling molecules via the pathways involved in the regulation of the cell cycle. This relationship between BAMBI and its involvement in the regulation of the cell cycle would provide a possibility that the BAMBI may be a new target for gene therapy.
[Show abstract][Hide abstract] ABSTRACT: Clear cell renal cell carcinoma (CRCC) is the most common malignant tumor of the kidney, and the clinical outcome of CRCC is related with the metastatic potential of CRCC. A significant proportion of metastatic CRCC remains incurable. Recently, immunotherapy against specific targets such as programmed death 1 (PD1) has been adapted for fatal cases of CRCC.
In this study, we aimed to evaluate the potential of tumor-infiltrating PD1-positive lymphocytes or FoxP3-positive regulatory T cells (Tregs) as predictors of the metastatic potential or prognosis of CRCC and investigate possible correlations with Epstein-Barr virus (EBV) infection in 199 cases of CRCC.
PD1 positivity, high Treg number, and EBV infection all predicted poor overall survival (OS) by univariate analysis. PD1 positivity and high Treg numbers were also significantly correlated with more distant metastatic relapse (DMR) and poor relapse-free survival (RFS) by univariate analysis. PD1 positivity and high Treg number were independent prognostic indicators for OS. In addition, PD1 positivity was an independent predictor of RFS and DMR. EBV infection was an independent predictor of OS of CRCC.
This study demonstrates that intratumoral infiltration of PD1-positive or FoxP3-positive lymphocytes can be used as significant prognostic indicators of CRCC and PD1 positivity could be very helpful in the prediction of latent distant metastasis of CRCCs. Therefore, evaluation of the infiltration of PD-positive cells or Tregs in CRCC may be useful diagnostic tools for the selection of patients who could benefit from PD1- or Treg-based immunotherapy.