JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Gain-of-function mutant p53 promotes cell growth and cancer cell metabolism via inhibition of AMPK activation.
Mol. Cell
PUBLISHED: 02-18-2014
Show Abstract
Hide Abstract
Many mutant p53 proteins (mutp53s) exert oncogenic gain-of-function (GOF) properties, but the mechanisms mediating these functions remain poorly defined. We show here that GOF mutp53s inhibit AMP-activated protein kinase (AMPK) signaling in head and neck cancer cells. Conversely, downregulation of GOF mutp53s enhances AMPK activation under energy stress, decreasing the activity of the anabolic factors acetyl-CoA carboxylase and ribosomal protein S6 and inhibiting aerobic glycolytic potential and invasive cell growth. Under conditions of energy stress, GOF mutp53s, but not wild-type p53, preferentially bind to the AMPK? subunit and inhibit AMPK activation. Given the importance of AMPK as an energy sensor and tumor suppressor that inhibits anabolic metabolism, our findings reveal that direct inhibition of AMPK activation is an important mechanism through which mutp53s can gain oncogenic function.
Related JoVE Video
A metabolomics-based approach for predicting stages of chronic kidney disease.
Biochem. Biophys. Res. Commun.
PUBLISHED: 02-04-2014
Show Abstract
Hide Abstract
Chronic kidney disease (CKD) is a major epidemiologic problem and a risk factor for cardiovascular events and cerebrovascular accidents. Because CKD shows irreversible progression, early diagnosis is desirable. Renal function can be evaluated by measuring creatinine-based estimated glomerular filtration rate (eGFR). This method, however, has low sensitivity during early phases of CKD. Cystatin C (CysC) may be a more sensitive predictor. Using a metabolomic method, we previously identified metabolites in CKD and hemodialysis patients. To develop a new index of renal hypofunction, plasma samples were collected from volunteers with and without CKD and metabolite concentrations were assayed by quantitative liquid chromatography/mass spectrometry. These results were used to construct a multivariate regression equation for an inverse of CysC-based eGFR, with eGFR and CKD stage calculated from concentrations of blood metabolites. This equation was able to predict CKD stages with 81.3% accuracy (range, 73.9-87.0% during 20 repeats). This procedure may become a novel method of identifying patients with early-stage CKD.
Related JoVE Video
Effect of a nitric oxide synthase inhibitor and a CXC chemokine receptor-4 antagonist on tumor growth and metastasis in a xenotransplanted mouse model of adenoid cystic carcinoma of the oral floor.
Int. J. Oncol.
PUBLISHED: 04-04-2013
Show Abstract
Hide Abstract
Nitric oxide (NO) is related to angiogenesis and tumor progression and chemokine receptor-4 (CXCR4) plays a central role in cell migration in metastasis and dissemination of cancer. The present study evaluated the effectiveness of a NOS inhibitor and a CXCR4 antagonist, given as single agents or in combination, in a xenotransplanted mouse model of adenoid cystic carcinoma (ACC) of the oral floor. A metastatic tumor (ACCIM) derived from a cervical metastatic lesion of human ACC that was transplantable in nude mice was used. ACCIM showed a high frequency of spontaneous metastasis to the lung when transplanted subcutaneously in nude mice. Mice with subcutaneous transplants of ACCIM were subdivided into six groups and intraperitoneally received one of the following treatments daily for 5 weeks: a) PBS (control), b) AMD3100 (CXCR4 antagonist), c) L-NAME (NOS inhibitor), d) 1400W (iNOS inhibitor), e) both AMD3100 and L-NAME (AMD3100+L-NAME) and f) both AMD3100 and 1400W (AMD3100+1400W). Tumor growth was evaluated during treatment and metastasis was assessed at 28 weeks. Single-agent treatment with AMD3100, L-NAME or 1400W inhibited tumor growth by 20.8, 26.5 and 54.5%, respectively. Combined treatment with AMD3100+L-NAME and AMD3100+1400W inhibited tumor growth remarkably by 48.0 and 50.2%, respectively. Immunohistochemical analysis revealed lower expression of CXCR4, iNOS and eNOS in tumor cells treated with AMD3100+L-NAME or AMD3100+1400W compared to control tumor cells and increased numbers of apoptotic tumor cells were demonstrated using the TUNEL method. CXCR4 expression decreased in 1400W-treated tumors using western blot analysis. When the effect of each agent on tumor-induced angiogenesis in tumor stroma was examined histologically, microvessel density was significantly lower in the groups treated with 1400W, AMD3100+L-NAME or AMD3100+1400W compared to the control, AMD3100 and L-NAME groups. Moreover, treatment with AMD3100 or 1400W markedly inhibited lung metastasis. Our results indicated that single-agent treatment with 1400W and combined treatment with AMD3100+L-NAME or AMD3100+1400W induced apoptosis and significantly inhibited tumor-induced angiogenesis and proliferation of ACCIM in vivo. Blockade of CXCR4 and iNOS was suggested to inhibit lung metastases from ACCIM. CXCR4 and iNOS may, thus, be important prognostic factors for long-term survival in ACC.
Related JoVE Video
Inferior alveolar nerve regeneration after bifocal distraction osteogenesis in dogs.
J. Oral Maxillofac. Surg.
PUBLISHED: 01-07-2013
Show Abstract
Hide Abstract
Bifocal distraction osteogenesis has been shown to be a reliable method for reconstructing missing bone segments. However, no reports have been published regarding inferior alveolar nerve regeneration during this procedure. We assumed that the nerve could regenerate with the bone regeneration during bifocal distraction, if the nerve had been saved at a mesial site of the transport disc. In the present study, we investigated that possibility in dogs.
Related JoVE Video
Antiproliferative effect of a novel mTOR inhibitor temsirolimus contributes to the prolonged survival of orthotopic esophageal cancer-bearing mice.
Cancer Biol. Ther.
PUBLISHED: 01-04-2013
Show Abstract
Hide Abstract
Esophageal squamous cell carcinoma (ESCC) remains one of the most aggressive cancers with poor prognosis regardless of a several reports that indicate a better therapeutic efficacy using some new chemotherapeutic agents. Recent drug development has contributed to an improved specificity to suppress mTOR activity by which many types of malignancies can be explosively progressed. Temsirolimus (CCI-779, TricelTM) is one of recently synthesized analogs of rapamycin and has provided better outcomes for patients with renal cell carcinoma. In this study, we experimentally evaluated an efficacy of targeting mTOR by temsirolimus for ESCC treatment, with an assessment of its survival advantage using an advanced ESCC animal model. First, we confirmed that the expression of phosphorylated mTOR was increased in 46 of 58 clinical ESCC tumor tissues (79.3%) and appeared to get strengthened with tumor progression. All of ESCC cell lines used in this study revealed an increase of mTOR phosphorylation, accompanied with the upregulation of hypoxia inducible factor-I ? (HIF-1?), one of the critical effectors regulated by mTOR. Temsirolimus treatment apparently suppressed the activation of mTOR and its downstream effectors, resulting in the reduced ability of ESCC cell proliferation. Finally, the weekly administration of temsirolimus significantly diminished the size of subcutaneous tumors (vehicle, 3261.6 ± 722.0; temsirolimus, 599.2 ± 122.9; p = 0.007) in nude mice and effectively prolonged orthotopic esophageal cancer-bearing mice (median survival periods: control, 31 d; temsirolimus, 43 d; p = 0.0024). These data suggests that targeting mTOR by temsirolimus may become a therapeutic alternative for esophageal cancer, with a contribution to a better outcome.
Related JoVE Video
Role of endothelin receptor signalling in squamous cell carcinoma.
Int. J. Oncol.
PUBLISHED: 09-12-2011
Show Abstract
Hide Abstract
Endothelin plays important roles in various physiological functions including vascular constriction. Recent studies reported that the endothelin receptors ETA and ETB are highly expressed in lung and skin tumor tissues. In contrast, there are few reports on endothelin signalling in the proliferation of head and neck cancer. We found that both ETA and ETB endothelin receptors were overexpressed in tumor cells of tongue cancer samples by immunohistochemistry. ETA and ETB were expressed in cultured lingual and esophageal squamous cell carcinoma (SCCs) cell lines. When both cultured cell lines were treated with an ETA selective antagonist (BQ123) or an ETB selective antagonist (BQ788), inhibition of cell growth was observed. Similar results were observed when SCCs were treated with specific siRNA for the suppression of ETA or ETB. Furthermore, inhibition of the mitogen-activated protein (MAP) kinase pathway by the treatments with ET receptor antagonists and siRNA was also observed. These results indicate that endothelin signalling may, in part, play important roles in cell growth in SCCs through the MAP kinase pathway.
Related JoVE Video
Tumor-selective, adenoviral-mediated GFP genetic labeling of human cancer in the live mouse reports future recurrence after resection.
Cell Cycle
PUBLISHED: 08-15-2011
Show Abstract
Hide Abstract
We have previously developed a telomerase-specific replicating adenovirus expressing GFP (OBP-401), which can selectively label tumors in vivo with GFP. Intraperitoneal (i.p.) injection of OBP-401 specifically labeled peritoneal tumors with GFP, enabling fluorescence visualization of the disseminated disease and real-time fluorescence surgical navigation. However, the technical problems with removing all cancer cells still remain, even with fluorescence-guided surgery. In this study, we report imaging of tumor recurrence after fluorescence-guided surgery of tumors labeled in vivo with the telomerase-dependent, GFP-containing adenovirus OBP-401.. Recurrent tumor nodules brightly expressed GFP, indicating that initial OBP-401-GFP labeling of peritoneal disease was genetically stable, such that proliferating residual cancer cells still express GFP. In situ tumor labeling with a genetic reporter has important advantages over antibody and other non-genetic labeling of tumors, since residual disease remains labeled during recurrence and can be further resected under fluorescence guidance.
Related JoVE Video
Inhibition of mTOR by temsirolimus contributes to prolonged survival of mice with pleural dissemination of non-small-cell lung cancer cells.
Cancer Sci.
PUBLISHED: 06-02-2011
Show Abstract
Hide Abstract
Temsirolimus (CCI-779), a recently synthesized analogue of rapamycin, specifically inhibits mTOR and has been approved for clinical use in renal cell carcinoma. Recent reports have indicated the growth inhibitory effect of temsirolimus in some cancers including non-small-cell lung carcinoma (NSCLC). In this study, we aimed to explore the potential therapeutic use of temsirolimus as a treatment for NSCLC. Using cultured NSCLC cells (A549, H1299, and H358), we determined the effect of temsirolimus on cell proliferation and its antitumor effects on subcutaneous tumors, as well as its contribution to the survival of mice having pleural dissemination of cancer cells, mimicking advanced NSCLC. Temsirolimus suppressed proliferation of NSCLC cells in a dose-dependent manner, with an IC(50) of <1 nM. Western blot analysis revealed that temsirolimus treatment specifically inhibited the phosphorylation of mTOR and its downstream effectors in 1 h, accompanied by an increased cell population in the G(0) /G(1) phase, but according to flow cytometry, the cell population did not increase in the sub-G(0) phase. When NSCLC subcutaneous tumor-bearing mice were treated with temsirolimus, tumor volume was significantly reduced (tumor volume on day 35: vehicle vs temsirolimus = 1239 vs 698 cm(3) ; P < 0.05). Furthermore, prolonged survival was observed in pleural disseminated tumor-bearing mice with temsirolimus treatment (median survival: vehicle vs temsirolimus = 53.5 vs 72.5 days; P < 0.05). These results suggest that temsirolimus could be useful for NSCLC treatment, due to its antiproliferative effect, and could be a potential treatment for advanced NSCLC, giving prolonged survival.
Related JoVE Video
Nationwide survey for bisphosphonate-related osteonecrosis of the jaws in Japan.
J. Oral Maxillofac. Surg.
PUBLISHED: 03-16-2011
Show Abstract
Hide Abstract
A nationwide retrospective cohort study was conducted by the Japanese Society of Oral and Maxillofacial Surgeons to assess the occurrence of bisphosphonate (BP)-related osteonecrosis of the jaws (BRONJ) during 2006 to 2008 and to elucidate the outcome and factors associated with remission of BRONJ.
Related JoVE Video
Effect of risedronate on osteoblast differentiation, expression of receptor activator of NF-?B ligand and apoptosis in mesenchymal stem cells.
Basic Clin. Pharmacol. Toxicol.
PUBLISHED: 03-16-2011
Show Abstract
Hide Abstract
Nitrogen-containing bisphosphonates (BPs) are antiresorptive drugs used for the treatment of metabolic bone diseases. Bone marrow stromal cells such as mesenchymal stem cells (MSCs) and MSC-derived osteoblasts that originate from MSCs are known to regulate osteoclast differentiation and activation via the expression of receptor activator of NF-?B ligand (RANKL). Although the effects of nitrogen-containing BPs on osteoclasts and osteoblasts have been well investigated, their effects in MSCs have not been clarified. In this study, we investigated the effects of risedronate (RIS), a nitrogen-containing BP, on osteoblast differentiation, RANKL expression and apoptosis in human and rat MSCs. RIS suppressed the formation of mineralized nodules and mRNA expression of differentiation marker genes such as bone sialoprotein and osteocalcin in MSC-derived osteoblasts. The RANKL expression induced by 1,25-(OH)(2) vitamin D(3) was not affected by RIS in human MSC-derived osteoblasts. In addition, treatment with high-concentration RIS induced chromatin condensation, an apoptosis feature, in MSCs. RIS-induced chromatin condensation was suppressed by a pan-caspase inhibitor zVAD-FMK and a cell-permeable isoprenoid analogue geranylgeraniol. These results indicate that RIS suppressed osteoblast differentiation and induced caspase- and isoprenoid depletion-dependent apoptosis and suggest that the antiresorptive effect of RIS is not mediated by a decrease in the RANKL expression in MSC-derived osteoblasts.
Related JoVE Video
Glutamine depletion induces murine neonatal melena with increased apoptosis of the intestinal epithelium.
World J. Gastroenterol.
PUBLISHED: 03-11-2011
Show Abstract
Hide Abstract
To investigate the possible biological outcome and effect of glutamine depletion in neonatal mice and rodent intestinal epithelial cells.
Related JoVE Video
Tubular adenomas with minor villous changes show molecular features characteristic of tubulovillous adenomas.
Am. J. Surg. Pathol.
PUBLISHED: 01-26-2011
Show Abstract
Hide Abstract
Advanced colorectal polyps are identified based on size ?10 mm, high-grade dysplasia, and/or villous histology. A diagnosis of tubular adenoma (TA) is recommended if villous change occupies <20% of the lesion, or tubulovillous adenoma (TVA) is recommended if there is 20% to 80% villosity. We hypothesized that even subtle villous changes (1% to 20%) would correlate with advanced molecular features. Two hundred sixty-nine colorectal adenomas were examined for KRAS and BRAF mutation and immunohistochemical staining of ?-catenin, O6-Methyl Guanine DNA Methyltransferase (MGMT), and p53. Adenomas were classified as TA1 (0% villosity, n=70), TA2 (1% to 20% villosity, n=81), or TVA (21% to 80% villosity, n=118). Clinical and molecular features were analyzed by univariate ?² and multivariate logistic regression. There was an incremental increase in KRAS mutation frequency with increasing villous compartment (17.9% TA1, 59.0% TA2, 78.4% TVA; P<0.001). MGMT was more frequently lost in TA2 (37.0%) than in TA1 (8.6%) (P<0.001) but did not differ from TVA (39.8%). p53 overexpression was more common in TA2 (38.3%) than in TA1 (10.0%) (P<0.001) but did not differ from TVA (32.2%). On multivariate analysis, TA2 adenomas were more likely to have a KRAS mutation [odds ratio (OR) 6.6, 95% confidence interval (CI), 3.0-14.2], MGMT loss (OR 6.2, 95% CI, 2.4-16.0), or p53 overexpression (OR 5.6, 95% CI, 2.3-13.7) than TA1. We have identified a subgroup of TAs based on subtle villous changes. These adenomas are more likely to show molecular features that are characteristic of TVAs than TAs. These data support the concept that any villous change may indicate increased malignant potential and may be useful to consider when assigning surveillance guidelines.
Related JoVE Video
Metabolomic analysis of human plasma from haemodialysis patients.
Eur. J. Clin. Invest.
PUBLISHED: 10-18-2010
Show Abstract
Hide Abstract
Urea and creatinine are widely used as biomarkers for disease. However, these parameters have been criticized as markers for several reasons. Thus, we conducted this study to identify novel biomarkers that can be used as alternatives to urea and creatinine to estimate the adequate dialysis dose by metabolomic analyses of plasma samples from patients undergoing haemodialysis.
Related JoVE Video
[Bacteria isolated from surgical infections and their susceptibilities to antimicrobial agents --special references to bacteria isolated between April 2008 and March 2009].
Jpn J Antibiot
PUBLISHED: 10-06-2010
Show Abstract
Hide Abstract
Bacteria isolated from infections in abdominal surgery during the period from April 2008 to March 2009 were investigated in a multicenter study in Japan, and the following results were obtained. In this series, 712 strains including 18 strains of Candida spp. were isolated from 173 (80.5%) of 215 patients with surgical infections. Three hundred and sixty-six strains were isolated from primary infections, and 346 strains were isolated from postoperative infections. From primary infections, anaerobic Gram-negative bacteria were predominant, followed by aerobic Gram-negative bacteria, while from postoperative infections aerobic Gram-positive bacteria were predominant, followed by anaerobic Gram-negative bacteria. Among aerobic Gram-positive bacteria, the isolation rate of Enterococcus spp. was highest, followed by Streptococcus spp., and Staphylococcus spp. in this order, from primary infections, while Enterococcus spp. was highest, followed by Staphylococcus spp. from postoperative infections. Among aerobic Gram-negative bacteria, Escherichia coli was the most predominantly isolated from primary infections, followed by Klebsiella pneumoniae and Pseudomonas aeruginosa, in this order, and from postoperative infections, P aeruginosa was most predominantly isolated, followed by E. coli, Enterobacter cloacae, and K. pneumoniae. Among anaerobic Gram-positive bacteria, the isolation rate of Eggerthella lenta was the highest from primary infections, followed by Parvimonas micra, Streptococcus constellatus and Gemella morbillorum, and from postoperative infections, E. lenta was most predominantly isolated. Among anaerobic Gram-negative bacteria, the isolation rate of Bacteroides fragilis was the highest from primary infections, followed by Bacteroides thetaiotaomicron, Bacteroides ovatus and Bilophila wadsworthia, and from postoperative infections, B. fragilis was most predominantly isolated, followed by B. thetaiotaomicron, B. wadsworthia and B. ovatus, in this order. In this series, we noticed no vancomycin-resistant methicillin-resistant S. aureus, and Enterococcus spp., nor multidrug-resistant P aeruginosa. We should carefully follow up B. wadsworthia which was resistant to various antibiotics, and also Bacteroides spp. which was resistant to many beta-lactam antibiotics.
Related JoVE Video
Preferential up-regulation of heparanase and cyclooxygenase-2 in carcinogenesis of Barretts oesophagus and intestinal-type gastric carcinoma.
Histopathology
PUBLISHED: 07-27-2010
Show Abstract
Hide Abstract
Metaplastic changes secondary to chronic inflammation at the gastro-oesophageal junction and at the pyloric antrum are recognized as the premalignant conditions of Barretts oesophageal adenocarcinoma and intestinal-type gastric carcinoma (GC), respectively. Heparanase (HPSE) and cyclooxygenase (COX)-2 have been proved to play critical roles in inflammation as well as in cancer. The aim was to examine the meaning of their expression in inflammation-related carcinogenesis.
Related JoVE Video
Clonal overgrowth of esophageal smooth muscle cells in diffuse leiomyomatosis-Alport syndrome caused by partial deletion in COL4A5 and COL4A6 genes.
Matrix Biol.
PUBLISHED: 06-29-2010
Show Abstract
Hide Abstract
This is a study of a patient who manifests all of the features of a diffuse leiomyomatosis-Alport syndrome (DL-ATS), and her two-year-old son who has already been diagnosed with Alport syndrome. Fourteen years ago, the patient underwent a partial esophageal resection followed by a replacement with jejunum. Recently, she underwent a surgical resection of the esophagus due to esophageal dysfunction. Genetic analyses of COL4A5 and COL4A6 on the X-chromosome were efficiently performed using the genomic DNA of her son. We have identified a novel deletion of 194-kb in length, encompassing COL4A5-COL4A6 promoters as well as nearly the entire large intron 1 of COL4A5 and intron 2 of COL4A6. To uncover the relationship of the esophagus-specific occurrence of the tumor and the expression of those genes, immunohistochemical analyses of type IV collagen ? chains were conducted in the non-affected individuals. The esophageal smooth muscle-specific expression of ?5(IV) and ?6(IV) chains in the gastrointestinal tract was observed. Moreover, CAG repeat analysis of the androgen receptor gene and an immunohistochemical analysis in the leiomyoma revealed clonal overgrowth of the cells which received X-inactivation on the non-affected allele. These results may suggest that the dominant effect was caused by the partial deletion of the esophageal smooth muscle-specific genes, COL4A5 and COL4A6.
Related JoVE Video
Isolation and propagation of a human CD133(-) colon tumor-derived cell line with tumorigenic and angiogenic properties.
Cell Transplant
PUBLISHED: 06-29-2010
Show Abstract
Hide Abstract
It has been proposed in human colorectal cancers (CRC) a minority subset of cancer cells within tumors able to initiate tumor growth, defined as cancer stem cells (CSC). Solid human primary colonic and its ovarian metastatic cancer tissues were collected from fresh surgical samples and subsequent xenografts were established in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. The resulting tumors were disaggregated into single-cell suspensions and a CD133(-) cell line (NANK) was newly established and analyzed by flow cytometry. Surface markers of progenitor cells were immunophenotypically analyzed, and expression of stem cell and cancer-related genes was characterized. Secreted angiogenesis-associated molecules were investigated by proteomic array technology. Finally, different numbers of NANK were implanted and their tumor-initiating properties were investigated in NOD/SCID mice. Intraperitoneal injection of NANK in NOD/SCID mice induced tumors with developing progressive peritoneal dissemination and ascites. NANK cells maintained a differentiated phenotype and reproduced the full morphologic and phenotypic heterogeneity of their parental lesions. Noticeably, NANK lacked the expression of conventional CSC markers CD133 and CD44, self-renewal genes Oct-4 and Nanog, but showed the expression of an important gastrointestinal development marker CDX-2 and BMI-1 that is essential in regulating the proliferative activity of normal and leukemic stem cells. In addition, NANK secreted high amounts of important angiogenic cytokines. These results provide a novel and extensive model in human CSC for studying the generation and maintenance of phenotypic heterogeneity in CRC.
Related JoVE Video
Characteristics of CD133(+) human colon cancer SW620 cells.
Cell Transplant
PUBLISHED: 06-29-2010
Show Abstract
Hide Abstract
Worldwide, colorectal cancer is the third most common type of cancer affecting both sexes. It has been proposed that a small subset of cancer cells (cancer stem cells) within each tumor is able to initiate tumor growth. In 2007, two research groups simultaneously identified a colon cancer stem cell population in human tumors by the use of CD133 expression. In the present study, we used a human colon cancer cell line, SW620, to analyze the cancer stem cell-like characteristics of CD133(+) cells in vitro and in vivo. In vitro, CD133(+) SW620 cells had a higher proliferative capacity, were more irradiation- and chemotherapy-resistant, and had a higher expression of ?-catenin compared with CD133(-) cells. Injections of either CD133(+) or CD133(-) cells into the skin or rectal mucosa of NOD/SCID mice led to tumors; however, injection of CD133(+) cells resulted in the formation of larger tumors. Tumors derived from injections of CD133(-) cells did not contain any CD133(+) cells, whereas tumors derived from injections of CD133(+) cells did contain CD133(+) cells, suggesting self-renewing capability. However, the proportion of CD133(+) cells in the newly formed tumors in vivo was lower than the proportion of CD133(+) cells in vitro. In conclusion, the human colon cancer cell line, SW620, contains both CD133(+) and CD133(-) phenotypes, and the CD133(+) phenotype has characteristics consistent with those of cancer stem cells.
Related JoVE Video
Bone repair using a hybrid scaffold of self-assembling peptide PuraMatrix and polyetheretherketone cage in rats.
Cell Transplant
PUBLISHED: 06-23-2010
Show Abstract
Hide Abstract
Self-assembling peptide scaffold (SAPS) is well known to have very good bone conduction properties. However, the intensity of SAPS is too weak to actually use it for a clinical bone regeneration. Therefore, we have produced a hybrid scaffold system that involves fabricating a cage from polyetheretherketone (PEEK) that has high intensity, filling the interior of this cage with SAPS, and then transplanted this hybrid scaffold to bone defects in rat femurs. After 28 days, soft X-ray radiographs and histological assessment revealed that good new bone formation was clearly observed in the defects transplanted the PEEK cage with SAPS, but not in the PEEK cage only. The PEEK cage maintained a form and osteoconduction ability of internal SAPS, and SAPS promoted bone formation inside the PEEK; therefore, each was in charge of intensity and bone regeneration separately. The present study suggests that hybrid scaffolds made from PEEK cages and SAPS can be useful tools for the regeneration of load-bearing bones, based on the idea that it should be possible to develop ideal bone filler materials by combining the strength of artificial bone with the bone regeneration and bone conduction properties of SAPS.
Related JoVE Video
Proton-coupled erythromycin antiport at rat blood-placenta barrier.
Drug Metab. Dispos.
PUBLISHED: 06-21-2010
Show Abstract
Hide Abstract
The aim of the present study was to characterize the mechanism of erythromycin transport at the blood-placenta barrier, using TR-TBT 18d-1 cells as a model of rat syncytiotrophoblasts. [(14)C]Erythromycin was taken up by TR-TBT 18d-1 cells with a Michaelis constant of 466 microM. Although the uptake was not dependent on extracellular Na(+) or Cl(-), it was increased at weakly alkaline pH. Significant overshoot of [(14)C]erythromycin uptake by placental brush-border membrane vesicles was observed in the presence of an outwardly directed proton gradient. These results indicate that erythromycin is transferred by the H(+)-coupled transport system in syncytiotrophoblasts. To address the physiological transport of erythromycin in rat placenta, fetal-to-maternal transport clearance was estimated by means of the single placental perfusion technique. Clearance of [(14)C]erythromycin was higher than that of [(14)C]inulin, a paracellular pathway marker, and was decreased by the addition of 5 mM erythromycin, indicating that saturable efflux system from fetus to mother is involved. The effect of various transporter inhibitors on [(14)C]erythromycin efflux from TR-TBT 18d-1 cells was evaluated. cyclosporin A, fumitremorgin C, and probenecid had no effect, whereas ethylisopropylamiloride, a specific inhibitor of Na(+)/H(+) exchangers (NHEs), was significantly inhibitory. These results suggest that erythromycin efflux transport at the rat blood-placenta barrier is mediated by an erythromycin/H(+) antiport system, driven by H(+) supplied by NHEs.
Related JoVE Video
Preclinical evaluation of differentially targeting dual virotherapy for human solid cancer.
Mol. Cancer Ther.
PUBLISHED: 05-25-2010
Show Abstract
Hide Abstract
Multimodal approaches combining drugs that differentially function is the most popular regimen for treating human cancer. Understanding the molecular mechanisms underlying the synergistic, potentiative, and antagonistic effects of drug combinations could facilitate the discovery of novel efficacious combinations. We previously showed that telomerase-specific replication-competent adenovirus (Telomelysin, OBP-301), in which the human telomerase reverse transcriptase promoter controls the adenoviral E1 gene expression, induces a selective antitumor effect in human cancer cells. Here, using E1-deleted replication-deficient adenovirus expressing the p53 tumor suppressor gene (Advexin, Ad-p53) and OBP-301, we investigate how these adenoviruses that kill tumor cells with different mechanisms could work in combination on human cancer. We found that E1-deficient Ad-p53 could kill cancer cells more efficiently in the presence of OBP-301 than Ad-p53 alone or OBP-301 alone, because Ad-p53 could become replication-competent by being supplied adenoviral E1 from coinfected OBP-301 in trans. Ad-p53 plus OBP-301 induced high levels of p53 protein expression without p21 induction, resulting in apoptotic cell death documented by active caspase-3 expression with a cytometric bead array and an increased subdiploid apoptotic fraction of the cell cycle. For in vivo evaluation, nude mice xenografted with human lung tumors received intratumoral injection of OBP-301 and/or Ad-p53. Analysis of the growth of implanted tumors showed an enhanced antitumor effect in combination therapy. Our data show that Ad-p53 in combination with OBP-301 induces not only oncolytic but also apoptotic cancer cell death and enhances antitumor activity in vitro and in vivo, providing potential merits as a multimodal treatment for human cancer.
Related JoVE Video
In vivo biological purging for lymph node metastasis of human colorectal cancer by telomerase-specific oncolytic virotherapy.
Ann. Surg.
PUBLISHED: 05-21-2010
Show Abstract
Hide Abstract
The aim of this study was to develop a less invasive way of targeting lymph node metastasis for the treatment of human gastrointestinal cancer. Lymphatic invasion is a major route for cancer cell dissemination, and adequate treatment of locoregional lymph nodes is required for curative treatment in patients with malignancies.
Related JoVE Video
RAD001 offers a therapeutic intervention through inhibition of mTOR as a potential strategy for esophageal cancer.
Oncol. Rep.
PUBLISHED: 03-06-2010
Show Abstract
Hide Abstract
Esophageal cancer is one of the most frequently occurring cancers in the world. Targeting therapy strategy of cancer with specific inhibitors is developing and has showed promising antitumor efficacy. It is known that mTOR is an important controller of cell growth. RAD001 (everolimus) is a specific inhibitor of mTOR that can block the mTOR signaling pathway. The purposes of this study was to explore the inhibitory effects of RAD001 on mTOR signaling and the mechanism of cell growth suppression by RAD001. We examined both the expression of mTOR, p70S6K and S6 in SEG-1 esophageal cancer cells and KOB-13 normal esophageal epithelial cells and the efficacy of RAD001 against SEG-1 esophageal cancer cells. mTOR, p70S6K and S6 were overexpressed in SEG-1 esophageal cancer cells compared with KOB-13 normal esophageal epithelial cells. SEG-1 esophageal cancer cells were sensitive to RAD001. The survival rate of the cells treated with RAD001 over 0.33 microM was significantly different compared with that of control (P<0.01). RAD001 inhibited the phosphorylation of mTOR (Ser2448) and S6 (Ser240/244) in different grades and the expressions of mTOR, p70S6K and S6. As a result, RAD001 induced a dose-dependent decrease in cell proliferation, G1/S arrest and damage of cell shape. Taken together, these data showed that RAD001 can inhibit mTOR signaling and proliferation in SEG-1 esophageal cancer cells in vitro. It offers a therapeutic intervention through inhibition of mTOR as a potential strategy for esophageal cancer.
Related JoVE Video
[A case of early poorly-differentiated neuroendocrine carcinoma of stomach].
Gan To Kagaku Ryoho
PUBLISHED: 02-16-2010
Show Abstract
Hide Abstract
A 45-year-old male was admitted to our hospital complaining of anemia. Gastric endoscopy showed a type IIa+IIc tumor at the anterior wall of the gastric angle. Based on the pathology of the biopsy specimen, poorly-differentiated adenocarcinoma was diagnosed. Computed tomography scans showed regional lymph node swelling. Distal gastrectomy with a D2 lymph node dissection was performed. On pathology, the tumor was immunohistochemically positive for chromogranin A and synaptophysin. The Ki67 index was 70%. The tumor was diagnosed as poorly-differentiated neuroendocrine carcinoma of the stomach. He was treated with S-1 and CPT-11. Neuroendocrine cell carcinoma of the stomach is rare and usually has a very poor prognosis. Thus, we are reporting this case of early poorly-differentiated neuroendocrine carcinoma of the stomach that was curatively resected and had 12-month survival without recurrence.
Related JoVE Video
The establishment of a new mouse model with orthotopic esophageal cancer showing the esophageal stricture.
Cancer Lett.
PUBLISHED: 01-09-2010
Show Abstract
Hide Abstract
We established a promising new experimental animal model with an orthotopic xenograft of esophageal cancer that successfully represents poor oral intake, a major clinical feature of esophageal cancer. The advantage of this model is that no surgical technique is required, only the injection of a cell suspension by a needle and syringe via the esophageal lumen from the mouth, which provides a high reproducibility of tumor implantation and a rapid progress of outcome. We propose that this model is useful to study cancer-related outcomes and for developing new therapies for esophageal cancer, and we expect it to make a contribution to clinical practice.
Related JoVE Video
Selective metastatic tumor labeling with green fluorescent protein and killing by systemic administration of telomerase-dependent adenoviruses.
Mol. Cancer Ther.
PUBLISHED: 11-03-2009
Show Abstract
Hide Abstract
We previously constructed telomerase-dependent, replication-selective adenoviruses OBP-301 (Telomelysin) and OBP-401 [Telomelysin-green fluorescent protein (GFP); TelomeScan], the replication of which is regulated by the human telomerase reverse transcriptase promoter. By intratumoral injection, these viruses could replicate within the primary tumor and subsequent lymph node metastasis. The aim of the present study was to evaluate the possibility of systemic administration of these telomerase-dependent adenoviruses. We assessed the antitumor efficacy of OBP-301 and the ability of OBP-401 to deliver GFP in hepatocellular carcinoma (HCC) and metastatic colon cancer nude mouse models. We showed that i.v. administration of OBP-301 significantly inhibited colon cancer liver metastases and orthotopically implanted HCC. Further, we showed that OBP-401 could visualize liver metastases by tumor-specific expression of the GFP gene after portal venous or i.v. administration. Thus, systemic administration of these adenoviral vectors should have clinical potential to treat and detect liver metastasis and HCC.
Related JoVE Video
[Bacteria isolated from surgical infections and their susceptibility to antimicrobial agents--special reference to bacteria isolated between April 2007 and March 2008].
Jpn J Antibiot
PUBLISHED: 10-29-2009
Show Abstract
Hide Abstract
Bacteria isolated from infections in abdominal surgery during the period from April 2007 to March 2008 were investigated in a multicenter study in Japan, and the following results were obtained. In this series, 707 strains including 24 strains of Candida spp. were isolated from 181 (79.0%) of 229 patients with surgical infections. Three hundred and ninety-five strains were isolated from primary infections, and 288 strains were isolated from postoperative infections. From primary infections, anaerobic Gram-negative bacteria were predominant, followed by aerobic Gram-negative bacteria, while from postoperative infections aerobic Gram-positive bacteria were predominant, followed by anaerobic Gram-negative bacteria. Among aerobic Gram-positive bacteria, the isolation rate of Enterococcus spp. was highest, followed by Streptococcus spp., and Staphylococcus spp. in this order, from primary infections, while Enterococcus spp. was highest, followed by Staphylococcus spp. from postoperative infections. Among aerobic Gram-negative bacteria, Escherichia coli was the most predominantly isolated from primary infections, followed by Klebsiella pneumoniae, Pseudomonas aeruginosa and Enterobacter cloacae, in this order, and from postoperative infections, P. aeruginosa was most predominantly isolated, followed by E. cloacae, E. coli and K. pneumoniae. Among anaerobic Gram-positive bacteria, the isolation rate of Parvimonas micra was the highest from primary infections, followed by Streptococcus constellatus and Gemella morbillorum, and from postoperative infections, Anaerococcus prevotii was most predominantly isolated. Among anaerobic Gram-negative bacteria, the isolation rate of both Bacteroides fragilis and Bilophila wadsworthia were the highest from primary infections, followed by Bacteroides thetaiotaomicron and Campylobacter gracilis, and from postoperative infections, B. thetaiotaomicron was most predominately isolated, followed by B. fragilis, Bacteroides caccae and B. wadsworthia in this order. In this series, we noticed no vancomycin-resistant Gram-positive cocci, nor multidrug-resistant P aeruginosa. There were nine strains of coagulase-negative Staphylococci which show higher MIC against teicoplanin more than 4 gg/mL, but all of them had good susceptibilities against various anti-MRSA antibiotics. We should carefully follow up B. wadsworthia which was resistant to various antibiotics, and also Bacteroides spp. which was resistant to many beta-lactam antibiotics.
Related JoVE Video
Neovascularization induced around an artificial device implanted in the abdomen by the use of gelatinized fibroblast growth factor 2.
Cell Transplant
PUBLISHED: 09-25-2009
Show Abstract
Hide Abstract
The development of a bioartificial pancreas (BAP) with immunoisolating fashion has been gaining attention as a new method for treating diabetes. We have been proceeding with the development of a bag-type BAP that can be easily implanted and that allows for the optional injection or rejection of cells at any time. If fibrosis develops around a BAP device, then the permeability of substances transmitted through a semipermeable membrane will decrease, thereby reducing the reactivity with glucose, so it is necessary for the material of the device to have an excellent histocompatibility. Furthermore, in order to improve the efficacy of BAP treatment, it is important to maintain an environment of ample blood flow around the device. We have created a bag-type device for BAP that is 20 x 20 mm in size and comprises two layers of membranes. We have used an EVAL membrane for the outer membrane of the two layers. The EVAL membrane is a semipermeable membrane with good insulin permeability, which functions as an immunoisolation membrane. The inner membrane consists of PAU-coated HD-PE (nonwoven material processed with polyaminourethan) and it is designed to function as a scaffold for cells. We used Lewis rats to determine whether the effectiveness of fibroblast growth factor 2 (bFGF) can be improved by concomitantly using bFGF with a capacity for blood vessel regeneration as well as bFGF immersed in a sheet of gelatin. We placed the BAP in the abdominal cavity and covered it with the greater omentum. We were able to significantly increase the blood flow and the number of new blood vessels in the tissue surrounding the BAP device by using gelatinized bFGF. There were only a few instances of fibrosis as a biological reaction to the EVAL membrane, and the infiltration of inflammatory cells was mild. There were no adverse effects related to implantation of the device. We confirmed in this study that the use of an implantable BAP device and bFGF allowed for a better blood flow around the BAP device. There were only minor instances of fibrosis and inflammation reaction around the BAP, thus indicating the BAP that we are currently developing to have an excellent histocompatibility.
Related JoVE Video
Hepatocytes from fibrotic liver possess high growth potential in vivo.
Cell Transplant
PUBLISHED: 09-25-2009
Show Abstract
Hide Abstract
Hepatocyte transplantation is effective for treating liver failure, but healthy donors as a source of hepatocytes are quite limited. The livers of patients with hepatic fibrosis could be an alternative source; however, few reports have examined the nature of hepatocytes from fibrotic livers (f-hepatocytes). In this study, we compared the growth of f-hepatocytes and hepatocytes from normal livers (n-hepatocytes). Hepatocytes were isolated from normal and CCl(4)-treated wild-type Fischer rats that express dipeptidyl dipeptidase IV (DPPIV) gene (DPPIV(+)). The n- and f-hepatocytes proliferated in culture at similar rates. Both types of hepatocytes were transplanted into DPPIV(-) mutant Fischer rats that had been treated with retrorsine to injure the liver and were partially hepatectomized (PHx) before transplantation. Both n- and f-DPPIV(+)-hepatocytes proliferated and formed colonies. The colony sizes of f-hepatocytes 21 days posttransplantation were approximately three times those of n-hepatocytes. The hepatocytes were analyzed using a fluorescence activated cell sorter (FACS). The FACS profile differed between f- and n-hepatocytes: f-hepatocytes were less granular, less autofluorescent, and smaller than n-hepatocytes. These characteristics of f-hepatocytes resembled those reported for small-sized n-hepatocytes (SHs), which are highly proliferative and preferentially express a unique set of 10 SH genes. However, f-hepatocytes preferentially expressed only five of the SH genes. The expression profile of f-hepatocytes was rather similar to that of proliferating n-hepatocytes in the regenerating liver after PHx. The f-hepatocytes were morphologically normal and did not show any preneoplastic phenotype. These normal and proliferative natures of f-hepatocytes in vivo suggest the fibrotic liver as a source of hepatocytes for transplantation.
Related JoVE Video
Analysis of fecal DNA methylation to detect gastrointestinal neoplasia.
J. Natl. Cancer Inst.
PUBLISHED: 08-21-2009
Show Abstract
Hide Abstract
The development of noninvasive screening tests is important to reduce mortality from gastrointestinal neoplasia. We sought to develop such a test by analysis of DNA methylation from exfoliated cancer cells in feces.
Related JoVE Video
In vivo internal tumor illumination by telomerase-dependent adenoviral GFP for precise surgical navigation.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 08-17-2009
Show Abstract
Hide Abstract
Cancer surgery requires the complete and precise identification of malignant tissue margins including the smallest disseminated lesions. Internal green fluorescent protein (GFP) fluorescence can intensely illuminate even single cells but requires GFP sequence transcription within the cell. Introducing and selectively activating the GFP gene in malignant tissue in vivo is made possible by the development of OBP-401, a telomerase-dependent, replication-competent adenovirus expressing GFP. This potentially powerful adjunct to surgical navigation was demonstrated in 2 nude mouse models that represent difficult surgical challenges--the resection of widely disseminated cancer. HCT-116, a model of intraperitoneal disseminated human colon cancer, was labeled by virus injection into the peritoneal cavity. A549, a model of pleural dissemination of human lung cancer, was labeled by virus administered into the pleural cavity. Only the malignant tissue fluoresced brightly in both models. In the intraperitoneal model of disseminated cancer, fluorescence-guided surgery enabled resection of all tumor nodules labeled with GFP by OBP-401. The data in this report suggest that adenoviral-GFP labeling tumors in patients can enable fluorescence-guided surgical navigation.
Related JoVE Video
Drug-regulatable cancer cell death induced by BID under control of the tissue-specific, lung cancer-targeted TTS promoter system.
Int. J. Cancer
PUBLISHED: 07-15-2009
Show Abstract
Hide Abstract
Gene therapy and virotherapy are among the approaches currently being used to treat lung cancer. The success of cancer gene therapy depends on treatments where different types of tumors can be selectively targeted and destroyed without affecting normal cells and tissue. Previously, we described a promoter system (TTS) that we designed that is specifically targeted to lung cancer cells but which does not affect other types of cells including stem cells. In our study, we have enhanced the utility of the TTS system by inserting the pro-apoptotic gene BH3 domain interacting death agonist (Bid) into the TTS promoter system (TTS/Bid) to create a drug regulatable lung cancer-specific gene therapy. A recombinant adenoviral vector was used to introduce TTS/Bid (Ad-TTS/Bid) into lung cancer cells. BID expression and apoptosis occurred in A549 pulmonary adenocarcinoma cells but little Bid expression or apoptosis occurred in MCF7 breast cancer cells or in normal human lung fibroblasts. The use of cisplatin enhanced the processing of full length BID to t-BID which significantly increased lung cancer-specific cell death. In in vivo experiments, intraperitonal injection of cisplatin enhanced the antitumor effects of the vector in a lung cancer xeno-graft mouse model. Moreover, dexamethasone effectively suppressed exogenous BID expression and the antitumor effect of Ad-TTS/Bid both in vitro and in vivo. Here, we describe the efficacy of the use of cisplatin and dexamethasone with the anti lung cancer promoter system (Ad-TTS/Bid) for a safe and effective gene therapy against advanced lung cancer.
Related JoVE Video
Efficient molecular screening of Lynch syndrome by specific 3 promoter methylation of the MLH1 or BRAF mutation in colorectal cancer with high-frequency microsatellite instability.
Oncol. Rep.
PUBLISHED: 05-09-2009
Show Abstract
Hide Abstract
It is sometimes difficult to diagnose Lynch syndrome by the simple but strict clinical criteria, or even by the definitive genetic testing for causative germline mutation of mismatch repair genes. Thus, some practical and efficient screening strategy to select highly possible Lynch syndrome patients is exceedingly desirable. We performed a comprehensive study to evaluate the methylation status of whole MLH1 promoter region by direct bisulfite sequencing of the entire MLH1 promoter regions on Lynch and non-Lynch colorectal cancers (CRCs). Then, we established a convenient assay to detect methylation in key CpG islands responsible for the silencing of MLH1 expression. We studied the methylation status of MLH1 as well as the CpG island methylator phenotype (CIMP) and immunohistochemical analysis of mismatch repair proteins on 16 cases of Lynch CRC and 19 cases of sporadic CRCs with high-frequency microsatellite instability (MSI-H). Sensitivity to detect Lynch syndrome by MLH1 (CCAAT) methylation was 88% and the specificity was 84%. Positive likelihood ratio (PLR) was 5.5 and negative likelihood ratio (NLR) was 0.15. Sensitivity by mutational analysis of BRAF was 100%, specificity was 84%, PLR was 6.3 and NLR was zero. By CIMP analysis; sensitivity was 88%, specificity was 79%, PLR was 4.2, and NLR was 0.16. BRAF mutation or MLH1 methylation analysis combined with MSI testing could be a good alternative to screen Lynch syndrome patients in a cost effective manner. Although the assay for CIMP status also showed acceptable sensitivity and specificity, it may not be practical because of its rather complicated assay.
Related JoVE Video
Advanced glycation end products enhance monocyte activation during human mixed lymphocyte reaction.
Clin. Immunol.
PUBLISHED: 04-27-2009
Show Abstract
Hide Abstract
Posttransplant diabetes mellitus (PTDM) is a frequent complication among transplant recipients. Ligation of advanced glycation end products (AGEs) with their receptor (RAGE) on monocytes/macrophages plays roles in the diabetes complications. The enhancement of adhesion molecule expression on monocytes/macrophages activates T-cells, leading to reduced allograft survival. We investigated the effect of four distinct AGE subtypes (AGE-2/AGE-3/AGE-4/AGE-5) on the expressions of intracellular adhesion molecule (ICAM)-1, B7.1, B7.2 and CD40 on monocytes, the production of interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha and the proliferation of T-cells during human mixed lymphocyte reaction (MLR). AGE-2 and AGE-3 selectively induced the adhesion molecule expression, cytokine production and T-cell proliferation. The AGE-induced up-regulation of adhesion molecule expression was involved in the cytokine production and T-cell proliferation. AGE-2 and AGE-3 up-regulated the expression of RAGE on monocytes; therefore, the AGEs may activate monocytes, leading to the up-regulation of adhesion molecule expression, cytokine production and T-cell proliferation.
Related JoVE Video
[A case report of FOLFOX treatment for primary duodenal carcinoma with multiple liver metastases].
Gan To Kagaku Ryoho
PUBLISHED: 04-22-2009
Show Abstract
Hide Abstract
We report a case of a woman in her sixties having primary duodenal carcinoma with multiple liver metastases who was treated with oxaliplatin in combination with infusional 5-fluorouracil/Leucovorin(FOLFOX regimen). After completing 2 courses of the chemotherapy, computed tomography showed a partial response without any severe adverse events. Now at 8 months, the PR stage has been maintained. So far, no standard therapeutic strategy for metastatic duodenal carcinoma has been developed. However, we suggest a FOLFOX regimen can be highly effective as a safe approach for continuously maintaining the quality of life of patients with this rare type of cancer.
Related JoVE Video
Advanced glycation end products subspecies-selectively induce adhesion molecule expression and cytokine production in human peripheral blood mononuclear cells.
J. Pharmacol. Exp. Ther.
PUBLISHED: 04-20-2009
Show Abstract
Hide Abstract
Advanced glycation end products (AGEs) are proteins or lipids that become glycated after exposure to diverse reducing sugars. Accumulation of AGEs induces diabetes complications. Microinflammation is a common major mechanism in the pathogenesis of diabetic vascular complications. Activation of monocytes/macrophages and T cells plays roles in the pathogenesis of atherosclerosis. The activation of T cells requires the enhanced expression of adhesion molecules on monocytes. AGEs activate monocytes by engaging the receptor for AGE (RAGE); however, little is known about the profile of agonist activity of diverse AGE moieties on monocytes. We investigated the effect of four distinct AGE subtypes (AGE-modified bovine serum albumin; AGE-2, AGE-3, AGE-4, and AGE-5) at concentrations ranging from 0.1 to 100 microg/ml on the expression of intercellular adhesion molecule-1, B7.1, B7.2, and CD40 on monocytes and its impact on the production of interferon-gamma and tumor necrosis factor-alpha in human peripheral blood mononuclear cells. Among the AGEs examined, AGE-2 and AGE-3 selectively induced adhesion molecule expression and cytokine production. Antagonism experiments using antibodies against adhesion molecules demonstrated that cell-to-cell interaction between monocytes and T/natural killer cells was involved in AGE-2- and AGE-3-induced cytokine production. AGE-2 and AGE-3 up-regulated the expression of RAGE on monocytes. The effects of AGE-2 and AGE-3 were inhibited by nuclear factor-kappaB and p38 mitogen-activated protein kinase inhibitors. These results indicated that AGE-2 and AGE-3 activated monocytes via RAGE, leading to the up-regulation of adhesion molecule expression and cytokine production.
Related JoVE Video
Immunohistochemical staining of liver grafts with a monoclonal antibody against HCV-Envelope 2 for recurrent hepatitis C after living donor liver transplantation.
J. Gastroenterol. Hepatol.
PUBLISHED: 04-17-2009
Show Abstract
Hide Abstract
We evaluated the expression of hepatitis C virus (HCV) antigen on liver grafts by immunohistochemical staining (IHS) using IG222 monoclonal antibody (mAb) against HCV-envelope 2 (E2).
Related JoVE Video
Involvement of focal adhesion kinase in the progression and prognosis of gastrointestinal stromal tumors.
Hum. Pathol.
PUBLISHED: 04-10-2009
Show Abstract
Hide Abstract
Gastrointestinal stromal tumors often express gene mutations to c-KIT and platelet-derived growth factor receptor-alpha, both of which result in constitutive activations of their signaling pathways that are quite essential for the proliferation and survival of tumor cells in most clinical gastrointestinal stromal tumors. Targeting these molecules provides a dramatic improvement to therapeutic strategy. To identify a new therapeutic target for gastrointestinal stromal tumor treatment, we focused on focal adhesion kinase, which is reported to be an up-regulated gene in clinical gastrointestinal stromal tumors, because so far no one has examined its expression status at the protein level. In this study, Western blot analysis revealed that all 10 of the examined gastrointestinal stromal tumor tissues strongly expressed focal adhesion kinase protein and that phosphorylated focal adhesion kinase was detected in 9 of them. Next, we assessed the expression status of focal adhesion kinase and phosphorylated focal adhesion kinase in 51 cases of gastrointestinal stromal tumor by immunohistochemistry. Positive stainings for focal adhesion kinase and phosphorylated focal adhesion kinase were confirmed in 44 (86.3%) and in 40 cases (78.4%) of the 51 gastrointestinal stromal tumors, respectively. We further found that the focal adhesion kinase-positive staining rate became higher along with the increased status of malignant behavior. Moreover, when the 51 gastrointestinal stromal tumors were divided into 2 groups based upon their focal adhesion kinase expression status, the 5-year overall survival of patients in the focal adhesion kinase-positive group (66.5%) was significantly poorer than that in the focal adhesion kinase-negative group (100%). These results indicate that the up-regulation of focal adhesion kinase protein may also contribute to the tumor progression of gastrointestinal stromal tumors and that focal adhesion kinase is a potential target for gastrointestinal stromal tumor treatment.
Related JoVE Video
Specific Removal of Monocytes from Peripheral Blood of Septic Patients by Polymyxin B-immobilized Filter Column.
Acta Med. Okayama
PUBLISHED: 02-28-2009
Show Abstract
Hide Abstract
Lipopolysaccharide (LPS) is one of the major causes of septic shock. The polymyxin B-immobilized filter column (PMX) was developed for the adsorption of endotoxin by direct hemoperfusion and has been used for the treatment of LPS-induced septic shock. In this study, we demonstrated that PMX also specifically bound monocytes from the peripheral blood leukocytes of septic patients by mean of an analysis of bound cells using immunocytochemical and electron microscopic techniques. The specific removal of monocytes from septic patients may produce beneficial effects by reducing the interaction between monocytes and functionally associated cells including vascular endothelial cells.
Related JoVE Video
Intramuscular transplantation of engineered hepatic tissue constructs corrects acute and chronic liver failure in mice.
J. Hepatol.
PUBLISHED: 02-01-2009
Show Abstract
Hide Abstract
Transplantation of isolated hepatocytes holds great promise as an alternative to whole organ liver transplantation. For treatment of liver failure, access to the portal circulation has significant risks and intrahepatic hepatocyte engraftment is poor. In advanced cirrhosis, transplantation into an extrahepatic site is necessary and intrasplenic engraftment is short-lived. Strategies that allow repeated extrahepatic infusion of hepatocytes could improve the efficacy and safety of hepatocyte transplantation for the treatment of liver failure.
Related JoVE Video
A novel antiangiogenic effect for telomerase-specific virotherapy through host immune system.
J. Immunol.
PUBLISHED: 01-22-2009
Show Abstract
Hide Abstract
Soluble factors in the tumor microenvironment may influence the process of angiogenesis; a process essential for the growth and progression of malignant tumors. In this study, we describe a novel antiangiogenic effect of conditional replication-selective adenovirus through the stimulation of host immune reaction. An attenuated adenovirus (OBP-301, Telomelysin), in which the human telomerase reverse transcriptase promoter element drives expression of E1 genes, could replicate in and cause selective lysis of cancer cells. Mixed lymphocyte-tumor cell culture demonstrated that OBP-301-infected cancer cells stimulated PBMC to produce IFN-gamma into the supernatants. When the supernatants were subjected to the assay of in vitro angiogenesis, the tube formation of HUVECs was inhibited more efficiently than recombinant IFN-gamma. Moreover, in vivo angiogenic assay using a membrane-diffusion chamber system s.c. transplanted in nu/nu mice showed that tumor cell-induced neovascularization was markedly reduced when the chambers contained the mixed lymphocyte-tumor cell culture supernatants. The growth of s.c. murine colon tumors in syngenic mice was significantly inhibited due to the reduced vascularity by intratumoral injection of OBP-301. The antitumor as well as antiangiogenic effects, however, were less apparent in SCID mice due to the lack of host immune responses. Our data suggest that OBP-301 seems to have antiangiogenic properties through the stimulation of host immune cells to produce endogenous antiangiogenic factors such as IFN-gamma.
Related JoVE Video
Establishment of a lymph node metastasis model from subcutaneous tumors of gastrointestinal stromal tumor model cells.
Oncol. Rep.
PUBLISHED: 01-17-2009
Show Abstract
Hide Abstract
There is currently no suitable animal model of metastasis using cultured human gastrointestinal stromal tumor cells even though the molecular mechanisms of c-KIT-mediated progression and metastasis should be clarified. Ba/F3 murine lymphocyte cells transduced with mutant c-KIT have been utilized to analyze some molecular mechanisms related to a constitutively activated c-KIT signaling and to assess the efficacy of molecular-targeted inhibitors. Using this cellular system, we coincidentally discovered the development of axillary and inguinal lymph node swelling three weeks after subcutaneous injection of Ba/F3 cells with c-KIT mutation into nude mice. Mutation-specific PCR detected c-KIT mutation in the swollen lymph nodes but not in unmetastasized normal lymph nodes, indicating that the lymph nodes contain tumor cells which should come from a primary subcutaneous tumor. Microscopic observation revealed tumor cells infiltrating through lymphatic follicles with Ki-67-positive staining to distinguish them from lymphocytes. The significance of this model is helpful to understand the molecular mechanisms of c-KIT-mediated metastasis and is useful for assessments of molecular therapeutics and in vivo imaging.
Related JoVE Video
Resistance to paclitaxel therapy is related with Bcl-2 expression through an estrogen receptor mediated pathway in breast cancer.
Int. J. Oncol.
PUBLISHED: 01-17-2009
Show Abstract
Hide Abstract
Taxanes are approved for the treatment of breast cancer that has spread to the lymph nodes, following surgery and doxorubicin containing chemotherapy. Taxanes have improved the survival of breast cancer patients, especially in estrogen receptor (ER) negative population in clinical settings. This time we examined the relationship between chemosensitivity to Taxanes and expresson of ERalpha in breast cancer cell lines. In vitro effects of paclitaxel in 4 ER-positive and 3 ER-negative breast cancer cell lines were investigated by MTT assay. We also investigated members of Bcl-2 family by Western blotting and RT-PCR to clarify their role in paclitaxel resistance both in ER-positive and in ER-negative cells. ER-negative cell lines were more sensitive to paclitaxel than ER-positive cells. ER-negative KPL-4 and ZR-75-30 cells, which were sensitive to paclitaxel, became resistant when they were treated with demethylation agent, 5-aza-2-deoxycytidine. Analysis of proapoptotic (Bax) and antiapoptotic (Bcl-2) molecules suggested that Bcl-2 is likely to have a role in the resistance of ER-positive cells. Bcl-2 expression was increased in a time-dependent manner after treatment of ER-positive cell lines with estrogen (E2). On the other hand, Bcl-2 was not detected in ER-negative cell lines. However, no significant difference was detected for Bax mRNA levels before and after E2 treatment in ER-positive and negative cell lines. Activation of ER gene expression in ER-negative KPL-4 cells by 5-aza-2-deoxycytidine resulted in up-regulation of Bcl-2 mRNA. To support our data, we examined paclitaxel sensitivity in ER-negative MDA-MB-231 and ER stable transfectant cells S30 and JM6. This experiment also showed ER-negative cells were sensitive to paclitaxel but ER-positive cells were resistant to it. These results suggest that ER influenced chemosensitivity to paclitaxel through regulation of Bcl-2 family and regulation of the pathway may be crucial to increase the efficacy of taxanes in ER-positive breast cancer.
Related JoVE Video
A simple biological imaging system for detecting viable human circulating tumor cells.
J. Clin. Invest.
PUBLISHED: 01-16-2009
Show Abstract
Hide Abstract
The presence of circulating tumor cells (CTCs) in the peripheral blood is associated with short survival, making the detection of CTCs clinically useful as a prognostic factor of disease outcome and/or a surrogate marker of treatment response. Recent technical advances in immunocytometric analysis and quantitative real-time PCR have made it possible to detect a few CTCs in the blood; however, there is no sensitive assay to specifically detect viable CTCs. Here, we report what we believe to be a new approach to visually detect live human CTCs among millions of peripheral blood leukocytes, using a telomerase-specific replication-selective adenovirus expressing GFP. First, we constructed a GFP-expressing attenuated adenovirus, in which the telomerase promoter regulates viral replication (OBP-401; TelomeScan). We then used OBP-401 to establish a simple ex vivo method that was able to detect viable human CTCs in the peripheral blood. The detection method involved a 3-step procedure, including the lysis of rbc, the subsequent addition of OBP-401 to the cell pellets, and an automated scan using fluorescence microscopy. OBP-401 infection increased the signal-to-background ratio as a tumor-specific probe, because the fluorescent signal was amplified only in viable, infected human tumor cells, by viral replication. This GFP-expressing virus-based method is remarkably simple and allows precise enumeration of CTCs.
Related JoVE Video
Inhibition of focal adhesion kinase as a potential therapeutic strategy for imatinib-resistant gastrointestinal stromal tumor.
Mol. Cancer Ther.
PUBLISHED: 01-14-2009
Show Abstract
Hide Abstract
Focal adhesion kinase (FAK) is often up-regulated in a variety of malignancies, including gastrointestinal stromal tumor (GIST), and its overexpression seems to be associated with tumor progressiveness and poor prognosis. GIST is well known to have a mutation to c-KIT; thus, a specific c-KIT inhibitor (imatinib) is recognized as the first-line chemotherapy for GIST, although a certain type of c-KIT mutation reveals a resistance to imatinib due to as yet uncertain molecular mechanisms. To assess the c-KIT mutation-related variation of cellular responses to imatinib, murine lymphocyte-derived Ba/F3 cells, which are stably transduced with different types of c-KIT mutation, were treated with either imatinib or a FAK inhibitor (TAE226), and their antitumor effects were determined in vitro and in vivo. A mutation at exon 11 (KITdel559-560) displayed a high sensitivity to imatinib, whereas that at exon 17 (KIT820Tyr) showed a significant resistance to imatinib in vitro and in vivo. KIT820Tyr cells appeared to maintain the activities of FAK and AKT under the imatinib treatment, suggesting that FAK might play a role in cell survival in imatinib-resistant cells. When FAK activity in those cells was inhibited by TAE226, cell growth was equally suppressed and the cells underwent apoptosis regardless of the c-KIT mutation types. Oral administration of TAE226 significantly diminished tumor growth in nude mice bearing KIT(820Tyr) xenografts. In summary, c-KIT mutation at exon 17 displayed a resistance to imatinib with maintained activations of FAK and subsequent survival signals. Targeting FAK could be a potential therapeutic strategy for imatinib-resistant GISTs.
Related JoVE Video
[A case of primary adenocarcinoma of small intestine responding to XELOX chemotherapy and leading to a partial metabolic response].
Gan To Kagaku Ryoho
Show Abstract
Hide Abstract
We report a case of adenocarcinoma of the small intestine responding to XELOX chemotherapy, leading to a partial metabolic response(PMR). The patient was a 58-year-old male with multiple peritoneal dissemination of adenocarcinoma of the small intestine. Chemotherapy with XELOX(L-OHP 130 mg/m² on day 1 , and capecitabine 1,000 mg/m2 on days 1-14)was performed. After 4 courses, a significant tumor reduction was obtained. This case suggests that chemotherapy with XELOX is a potential regimen for small intestinal adenocarcinoma.
Related JoVE Video
Differential expression of aquaporin 5 and aquaporin 3 in squamous cell carcinoma and adenoid cystic carcinoma.
Int. J. Oncol.
Show Abstract
Hide Abstract
Aquaporins (AQPs) are a membrane protein family involved in the selective transport of water across cell membranes. Recent studies have reported the expression of AQP5 in several tumor types such as gastric, pulmonary, ovarian, pancreatic and colorectal cancer. We have previously reported the expression on tumor cells and the important role of AQP3 on cell growth in tongue cancer. However, little is known about the expression and precise role of AQP5 on squamous cell carcinoma (SCC) of the tongue. We investigated the expression of AQP5 and AQP3 in human oral SCC and adenoid cystic carcinoma (ACC). Overexpression of both AQP5 and AQP3 were immunohistochemically observed on tumor cells in SCC, whereas ACC cells were faintly stained with those antibodies against AQPs. Treatment with pan-AQP inhibitor or specific AQP5-siRNA showed inhibition of cell growth in SCC cell lines via the inhibition of integrins and the mitogen-activated protein kinase pathway. AQPs play important roles in cell growth in SCC rather than ACC.
Related JoVE Video
Steroid-free living donor liver transplantation for HCV--a multicenter prospective cohort study in Japan.
Clin Transplant
Show Abstract
Hide Abstract
This prospective, non-randomized, multicenter cohort study analyzed the safety and efficacy of a steroid-free immunosuppressive (IS) protocol for hepatitis C virus (HCV)-positive living donor liver transplant (LDLT) recipients in Japan. Of 68 patients enrolled from 13 transplant centers, 56 fulfilled the inclusion/exclusion criteria; 27 were assigned the steroid-free IS protocol (Fr group) and 29 the traditional steroid-containing IS protocol (St group). Serum HCV RNA levels increased over time and were higher in the St group until postoperative day 90 (POD 14, p=0.013). Preemptive anti-HCV therapy was started in a higher percentage of recipients (59.3%) in the Fr group than in the St group (31.0%, p=0.031), mainly due to early HCV recurrence. The incidence of HCV recurrence at one yr was lower in the Fr group (22.2%) than in the St group (41.4%; p=0.066). The incidence of acute cellular rejection was similar between groups. New onset diabetes after transplant, cytomegalovirus infection, and renal dysfunction were significantly less frequent in the Fr group than in the St group (p=0.022, p<0.0001, p=0.012, respectively). The steroid-free IS protocol safely reduced postoperative morbidity and effectively suppressed both the HCV viral load in the early post-transplant period and HCV recurrence in HCV-positive LDLT recipients.
Related JoVE Video
[A case of resected rectal cancer with hepatic node and multiple liver metastases effectively treated by preoperative modified FOLFOX6 and sLV5FU2 chemotherapy].
Gan To Kagaku Ryoho
Show Abstract
Hide Abstract
A 55-year-old male had complained of melena.Colonoscopy revealed a type 2 tumor at the rectum.CT demonstrated hepatic lymph nodes and multiple liver metastases(stage IV).Low anterior resection was performed(tub2, RsRa, circ, type 2, pSS, pN1, sH3, cHN1, sP0, cM0: fstage IV).The patient was treated with mFOLFOX6 and sLV5FU2 after operation.CT revealed a partial response after 14 courses of systemic chemotherapy.sLV5 FU2 therapy was converted to capecitabine because he experienced bone marrow suppression.CT showed that the liver metastases had enlarged but the hepatic lymph nodes disappeared.Right portal vein embolization was performed.After 4 weeks, right hepatectomy and hepatic lymph node dissection were performed.Preoperative chemotherapy with mFOLFOX6 seems beneficial as a neoadjuvant chemotherapy for hepatic lymph node-positive advanced colorectal cancer.
Related JoVE Video
Impact of CYP3A5*3 on Plasma Exposure and Urinary Excretion of Fentanyl and Norfentanyl in the Early Postsurgical Period.
Ther Drug Monit
Show Abstract
Hide Abstract
The pharmacokinetic characteristics of intravenous fentanyl have not been fully clarified in the early postsurgical period. The aim of this study was to evaluate the plasma exposure and urinary excretion of fentanyl and norfentanyl according to cytochrome P450 (CYP) 3A5 genetic polymorphism.
Related JoVE Video
Exploration of novel predictive markers in rat plasma of the early stages of chronic renal failure.
Anal Bioanal Chem
Show Abstract
Hide Abstract
To identify blood markers for early stages of chronic kidney disease (CKD), blood samples were collected from rats with adenine-induced CKD over 28 days. Plasma samples were subjected to metabolomic profiling by liquid chromatography-mass spectrometry, followed by multivariate analyses. In addition to already-identified uremic toxins, we found that plasma concentrations of N6-succinyl adenosine, lysophosphatidylethanolamine 20:4, and glycocholic acid were altered, and that these changes during early CKD were more sensitive markers than creatinine concentration, a universal indicator of renal dysfunction. Moreover, the increase in plasma indoxyl sulfate concentration occurred earlier than increases in phenyl sulfate and p-cresol sulfate. These novel metabolites may serve as biomarkers in identifying early stage CKD.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.