Knowledge and Awareness of Human Papillomavirus Vaccination and Cervical Cancer among Men and Women in Japan: A Questionnaire Survey.

BACKGROUND: In many advanced countries other than Japan, the incidence and mortality rates of cervical cancer, which is mainly caused by the human papillomavirus (HPV) infection, are decreasing probably due to the high rate of HPV vaccination and cervical cancer screening. In Japan, these rates are on the rise owing to the stagnation of vaccination and low screening rate. To improve these situations, active promotion of HPV vaccination and screening is required. As a preliminary stage, we investigated perceptions regarding cervical cancer and HPV vaccines among Japanese men and women and examined the difference in perceptions by sex. METHODS: This was a prospective cross-sectional questionnaire survey targeting Sojo University students and working adults. University students were targeted before learning about cervical cancer. Working adults were recruited on the basis of information from the Health Promotion of Health and Welfare Department of Kumamoto Prefectural Government in Japan and from companies via student organizations promoting cancer prevention. We surveyed respondents' knowledge and awareness about HPV vaccination and cervical cancer and performed logistic regression analysis to compare the results between men and women. RESULT: A total of 557 completed questionnaires (205 men and 352 women) were analyzed. Women had high levels of knowledge and awareness about HPV vaccination and cervical cancer compared with men. However, 70% of women surveyed had never been screened for cervical cancer. CONCLUSION: A total of 557 completed questionnaires (205 men and 352 women) were analyzed. Women had high levels of knowledge and awareness about HPV vaccination and cervical cancer compared with men. However, among surveyed women, the degree of knowledge and awareness was lower than that among women in other countries with established HPV vaccination programs. Furthermore, 70% of women surveyed had never been screened for cervical cancer.

Recent topics and advanced therapies in chronic granulomatous disease.

Chronic granulomatous disease (CGD) is a primary immunodeficiency characterized by the inability of phagocytes to produce reactive oxygen species (ROS) owing to a defect in any of the five components (CYBB/gp91phox, CYBA/p22phox, NCF1/p47phox, NCF2/p67phox, and NCF4/p40phox) and a concomitant regulatory component of Rac1/2 and CYBC1/Eros of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. Patients with CGD are at an increased risk of life-threatening infections caused by catalase-positive bacteria and fungi and of inflammatory complications such as CGD colitis. Antimicrobial and azole antifungal prophylaxes have considerably reduced the incidence and severity of bacterial and improved fungal infections and overall survival. CGD studies have revealed the precise epidemiology and role of NADPH oxidase in innate immunity which has led to a new understanding of the importance of phagocyte oxygen metabolism in various host-defense systems and the fields leading to cell death processes. Moreover, ROS plays central roles in the determination of cell fate as secondary messengers and by modifying of various signaling molecules. According to this increasing knowledge about the effects of ROS on the inflammasomal system, immunomodulatory treatments, such as IFN-γ and anti-IL-1 antibodies, have been established. This review covers the current topics in CGD and the relationship between ROS and ROS-mediated pathophysiological phenomena. In addition to the shirt summary of hematopoietic stem cell transplantation and gene therapy, we introduce a novel ROS-producing enzyme replacement therapy using PEG-fDAO to compensate for NADPH oxidase deficiency.

Treatment with Polyethylene Glycol-Conjugated Fungal D-Amino Acid Oxidase Reduces Lung Inflammation in a Mouse Model of Chronic Granulomatous Disease.

Chronic granulomatous disease (CGD) is a primary immunodeficiency wherein phagocytes are unable to produce reactive oxygen species (ROS) owing to a defect in the nicotinamide adenine dinucleotide phosphate oxidase (NADPH) complex. Patients with CGD experience bacterial and fungal infections and excessive inflammatory disorders. Bone marrow transplantation and gene therapy are theoretically curative; however, residual pathogenic components cause inflammation and/or organic damage in patients. Moreover, antibiotic treatments may not help in preventing excessive inflammation due to the residual presence of fungal cell wall ß-glucan. Thus, better treatment strategies against CGD are urgently required. Polyethylene glycol-conjugated recombinant porcine D-amino acid oxidase (PEG-pDAO) supplies ROS to defective NADPH oxidase in neutrophils of patients with CGD, following which the neutrophils regain bactericidal activity in vitro. In this study, we employed an in vivo nonviable Candida albicans (nCA)-induced lung inflammation model of gp91-phox knockout CGD mice and supplied novel PEG conjugates of Fusarium spp. D-amino acid oxidase (PEG-fDAO), as it exhibits higher enzyme activity than PEG-pDAO. The body weight, lung weight, and lung pathology were evaluated using three experimental strategies with the in vivo lung inflammation model to test the efficacy of the ROS-generating enzyme replacement therapy with PEG-fDAO. The lung weight and pathological findings suggest the condition was ameliorated by administration PEG-fDAO, followed by intraperitoneal injection of D-phenylalanine or D-proline. Although a more precise protocol is essential, these data reveal the targeted delivery of PEG-fDAO to the nCA-induced inflammation site and show that PEG-fDAO can be used to treat inflammation in CGD in vivo.

Consistency between patients and families in recognizing cancer chemotherapy side effects: A questionnaire survey.

BACKGROUND: Although the side effects of cancer chemotherapy impair a patient's quality of life, family members' awareness of side effects may relieve patient anxiety and distress. AIM: We investigated whether patients and their families were consistent in recognizing the occurrence and severity of symptomatic side effects of chemotherapy treatment for cancer. METHODS AND RESULTS: This was a prospective observational study. We administered a questionnaire survey to patients and family members to assess the frequency of occurrence (1: never, 2: almost never, 3: sometimes, 4: frequently, 5: almost always, 6: unknown) and the degree of severity (1: mild, 2: moderate, 3: severe, 4: extremely severe, 5: unknown) of physical and psychological symptoms associated with cancer chemotherapy. Weighted Kappa and Cramer coefficients were used to assess consistency between the two groups. We surveyed 20 pairs of patients (5 men, 15 women) and their families (10 men, 10 women); 17 pairs lived together. The median age was 65.5 years (interquartile [IQR], 58.75, 69.25) for patients and 61.00 years (IQR, 47.25, 71.25) for family members. Of patients, 17 had solid cancer, and three had leukemia. Family members mostly recognized objectively visible symptoms such as hair loss and development of spots and keratinization. However, it was difficult for families to detect invisible subjective symptoms such as weakness, dysesthesia, depressed mood, and unarticulated anxiety. CONCLUSIONS: The results indicated that recognition of invisible subjective symptoms in patients undergoing chemotherapy was difficult even for family members. Therefore, a multidisciplinary approach in which various medical professionals actively communicate with both patients and families is important. Information sharing in collaboration with patients and families could increase understanding of the patient's condition and optimize patient care.

A novel polysaccharide derived from algae extract inhibits cancer progression via JNK, not via the p38 MAPK signaling pathway.

Cancer has long been one of the most malignant diseases worldwide. Processes in cancer cells are often mediated by Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38 MAPK) and other signaling pathways. Traditional therapies are often problematic. Recently, a novel polysaccharide derived from algae extract was investigated due to the increasing interest in biological activities of compounds from marine organisms. The effect of this novel polysaccharide on human MKN45 gastric carcinoma cells was determined previously. The current aimed to determine whether the polysaccharide affects other types of cancer, and the deeper mechanisms involved in the process. Human MCF-7 breast cancer cells were used to investigate the novel polysaccharide for its role in the cell growth and migration, and determine the mechanisms affected. MTT assay, nuclear staining and fluorescence activated cell sorting analysis demonstrated that the novel polysaccharide reduced the viability of MCF-7 cells by inducing cell apoptosis and arresting the cells at G2/M phase. Results of western blot analysis demonstrated that phosphorylation of JNK and expression of p53, caspase-9 and caspase-3 were upregulated in the polysaccharide-treated MCF-7 cells. SP600125, an inhibitor of JNK, maintained MCF-7 cell viability, prevented cell apoptosis and cycle arrest, and downregulated the polysaccharide-induced protein phosphorylation/expression. However, a migration assay demonstrated that the novel polysaccharide did not change the migration of MCF-7 cells, as well as the expression of p38 MAPK, and matrix metalloproteinase-9 and -2. Taken together, the current study demonstrated that the novel polysaccharide suppressed cancer cell growth, induced cancer cell apoptosis and cell cycle arrest via JNK signaling, but had no effect on cancer cell migration and p38 MAPK signaling.

Functional validation and expression analysis of myotubes converted from skin fibroblasts using a simple direct reprogramming strategy.

Previously, we reported that MyoD, a master gene for myogenic cells, could efficiently convert primary skin fibroblasts into myoblasts and myotubes, thereby effecting direct reprogramming. In this study, we further demonstrated that MyoD-expressing primary fibroblasts displayed rapid movement in culture, with a movement velocity that was significantly faster, almost four times, than mouse primary myoblasts. MyoD-transduced cells obtained the characteristics of Ca2 + release and electrically-stimulated contraction, which was comparable to C2C12 myotubes, suggesting that the essential features of muscle were observed in the transduced cells. Furthermore, the ability to fuse to the host myoblasts means that gene transfer from MyoD-transduced cells to host muscle cells could be obtained by cell fusion. In comparison with the iPS method (indirect reprogramming), our transduction method has a low risk for tumorigenesis and carcinogenesis because the starting cells are fibroblasts and the transduced cells are myoblasts, both normal and mortal cells. Accordingly, MyoD transduction of human skin fibroblasts using the adenoviral vector is a simple, inexpensive and promising candidate as a new cell transplantation therapy for patients with muscular disorders.

A novel polysaccharide derived from algae extract induces apoptosis and cell cycle arrest in human gastric carcinoma MKN45 cells via ROS/JNK signaling pathway.

In recent years, interest in biological activities of compounds from marine organisms has intensified. Cancer is the most principal enemy for human life and health. For the first time, to the best of our knowledge, we investigated a novel algae-derived polysaccharide for its role in inducing apoptosis and cell cycle arrest in human gastric carcinoma MKN45 cells. We found that the novel polysaccharide suppressed MKN45 cell proliferation, induced cell apoptosis and arrested the cells at G2/M phase. Furthermore, we observed that the generation of reactive oxygen species (ROS) and the phosphorylation of Jun N-terminal kinase (JNK), p53, caspase-9 and -3 were induced in the polysaccharide-treated MKN45 cells. In addition, pretreatment with N-acetyl-cysteine (NAC) and SP600125, the inhibitor of ROS and JNK, induced MKN45 cell proliferation, prevented the cell apoptosis and released the cells from cycle arrest. Finally, we found that pretreatment with NAC prevented the JNK, p53, caspase-9 and -3 protein phosphorylation induced by the polysaccharide, however, pretreatment with SP600125 did not affect the generation of ROS, suggesting that ROS is upstream of JNK. Taken together, the novel polysaccharide induced cancer cell apoptosis and arrested cell cycle via ROS/JNK signaling pathway.

Silencing heme oxygenase-1 gene expression in retinal pigment epithelial cells inhibits proliferation, migration and tube formation of cocultured endothelial cells.

Heme oxygenase-1 (HO-1) plays an important role in the vasculature and in the angiogenesis of tumors, wounds and other environments. Retinal pigment epithelial (RPE) cells and choroidal endothelial cells (CECs) are the main cells involved in choroidal neovascularization (CNV), a process in which hypoxia plays an important role. Our aim was to evaluate the role of human RPE-cell HO-1 in the angiogenic activities of cocultured endothelial cells under hypoxia. Small interfering RNA (siRNA) for HO-1 was transfected into human RPE cell line ARPE-19, and zinc protoporphyrin (ZnPP) was used to inhibit HO-1 activity. Knockdown of HO-1 expression and inhibition of HO-1 activity resulted in potent reduction of the expression of vascular endothelial growth factor (VEGF) under hypoxia. Furthermore, knockdown of HO-1 suppressed the proliferation, migration and tube formation of cocultured endothelial cells. These findings indicated that HO-1 might have an angiogenic effect in CNV through modulation of VEGF expression and might be a potential target for treating CNV.

A novel polysaccharide compound derived from algae extracts protects retinal pigment epithelial cells from high glucose-induced oxidative damage in vitro.

Diabetic retinopathy is a common complication of diabetes mellitus (DM). The oxidative damage inflicted on retinal pigment epithelial (RPE) cells by high glucose closely approximates the molecular basis for the loss of vision associated with this disease. We investigate a novel algae-derived polysaccharide compound for its role in protecting ARPE-19 cells from high glucose-induced oxidative damage. ARPE-19 cells were cultured for 4 d with normal concentration of D-glucose, and exposed to either normal or high concentrations of D-glucose in the presence or absence of the polysaccharide compound at variety of concentrations for another 48 h. Taurine was used as a positive control. Activity of super oxide dismutase (SOD) and concentration of glutathione (GSH) were measured as well as cytotoxicity of high glucose and the polysaccharide compound. To analyse cellular damage by high glucose, activation of Annexin V and p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) were examined. Our results showed that a significant cellular damage on ARPE-19 cells after 48 h treatment with high glucose, accompanied by a decrease in SOD activity and GSH concentration; high glucose also caused ARPE-19 cell apoptosis and activation of p38MAPK and ERK. As the non-toxic polysaccharide compound protected ARPE-19 cells from high glucose-induced cellular damage, the compound recovered SOD activity and concentration of GSH in the cells. The compound also abrogated the cell apoptosis and activation of p38MAPK and ERK. Therefore, the polysaccharide compound derived from algae extracts could be unique candidate for a new class of anti-DM and anti-oxidative damage.

Inhibition of high glucose-induced VEGF and ICAM-1 expression in human retinal pigment epithelium cells by targeting ILK with small interference RNA.

The aim of this study was to investigate the change of Integrin-linked kinase (ILK) expression of human retinal pigment epithelium (RPE) cells in response to high glucose, and the effect of targeting ILK with small interference RNA (siRNA) on the high glucose-induced expression of vascular endothelial growth factor (VEGF) and intercellular adhesion molecule-1 (ICAM-1). The ILK mRNA and protein expression in human RPE cells were analyzed with RT-PCR and western blot after exposure to 5.5, 30, 40, 50 mM glucose, or 5.5 mM glucose+45.5 mM mannitol for 48 h. The expression of VEGF and ICAM-1 was also determined. Cells were treated with ILK siRNA, to determine the effect of ILK on VEGF and ICAM-1 expression following treatment with high glucose. High concentrations of glucose significantly up-regulated ILK mRNA and protein expression, and the ILK expression increased along with the glucose concentration. The changes of VEGF and ICAM-1 expression were similar to that of ILK expression. Knocking down ILK gene expression with siRNA inhibited the elevation of VEGF and ICAM-1 induced by high glucose treatment. These results suggested that ILK was involved in the response of RPE cells to high glucose and may therefore play a role in the pathogenesis of diabetic ophthalmology.

Protective effect of canolol from oxidative stress-induced cell damage in ARPE-19 cells via an ERK mediated antioxidative pathway.

PURPOSE: Oxidative stress damage to retinal pigment epithelial (RPE) cells is thought to play a critical role in the pathogenesis of age-related macular degeneration (AMD). This study was conducted to investigate the protective effect of canolol against oxidative stress-induced cell death in ARPE-19 cells and its underlying mechanism. METHODS: ARPE-19 cells, a human retinal pigment epithelial cell line, were subjected to oxidative stress with 150 µM t-butyl hydroxide (t-BH) in the presence/absence of canolol in different concentrations. Cell viabilities were monitored by a 3-(4, 5-dimethylthiazol-2-yl)-2, 5 diphenyl tetrazolium bromide (MTT) assay. The apoptosis was measured by flow cytometry using Annexin V-FITC and PI staining and intracellular reactive oxygen species (ROS) levels were measured by a fluorescence spectrophotometer. Gene expression of NF-E2-related factor (Nrf-2), heme oxygenase-1 (HO-1), catalase and glutathione S-transferase-pi (GST-pi) were measured by a reverse transcription polymerase chain reaction (RT-PCR) assay. Activation of the extracellular signal regulated kinase (ERK) protein was evaluated by western blot analysis. RESULTS: Canolol showed relatively high safety for ARPE-19 cells and recovered the cell death caused by t-BH dose-dependently at a concentration of 50-200 µM. Canolol also reduced t-BH-induced intracellular ROS generation and thus protected ARPE-19 cells from cell apoptosis. HO-1, catalase, GST-pi, and Nrf-2 were elevated in ARPE-19 cells after treatment with different concentrations of canolol for 24 h. Finally, canolol was found to activate extracellular signal regulated kinase (ERK) phosphorylation in ARPE-19 cells under the condition, with or without t-BH. CONCLUSIONS: Canolol protected ARPE-19 cells from t-BH-induced oxidative damage and the protective mechanism was associated, at least partly, with the upregulation (activation) of antioxidative enzymes, probably through an ERK mediated pathway. This suggests that canolol offers a remarkable protective effect against oxidative damage of RPE cells and may have a therapeutic effect on AMD and other oxidative stress-related retinal diseases.

The stimulatory activities of polysaccharide compounds derived from algae extracts on insulin secretion in vitro.

We prepared two series of polysaccharide compounds derived from algae extracts and investigated their stimulatory activity on insulin secretion in vitro using the rat pancreatic cell line, RIN-5F. Several of the compounds exhibited significant stimulatory activity in a dose-dependent manner without apparent cytotoxicity at concentrations above 10 microM. Glybenclamide, a commonly prescribed sulfonylurea (SU) against diabetes mellitus type II, was used as a positive control and showed moderate cytotoxicity in the cell culture assay system. Amylin (IAPP; islet amyloid polypeptide), an inhibitor for glybenclamide, did not inhibit the activity of the isolated compounds, suggesting that they act through a mechanism(s) different from glybenclamide. Algae-derived extracts could be candidates for a new class of anti-diabetic drugs.

Efficient conversion of ES cells into myogenic lineage using the gene-inducible system.

We established genetically engineered ES (ZHTc6-MyoD) cells that harbor a tetracycline-regulated expression vector encoding myogenic transcriptional factor MyoD, for the therapy of muscle diseases, especially Duchenne muscular dystrophy (DMD). Almost all the ZHTc6-MyoD cells were induced into muscle lineage after removal of tetracycline. The undifferentiated ZHTc6-MyoD cells are Sca-1+ and c-kit+, but CD34-, all well-known markers for mouse hematopoietic stem cells. In addition, they are able to maintain themselves in the undifferentiated state, even after one month of culture. Therefore, it is possible to obtain a large quantity of ZHTc6-MyoD cells in the undifferentiated state that maintain the potential to differentiate only into muscle lineage. Additionally, at two weeks post-injection of these cells into muscle of mdx, a model mouse of DMD, clusters of dystrophin-positive myofibers were observed at the injection site. Therefore, ES cells have considerable therapeutic potential for treating muscle diseases.

Adenoviral mediated MyoD gene transfer into fibroblasts: myogenic disease diagnosis.

MyoD, a master regulatory gene for myogenesis, converts mesoderm derived cells to the skeletal muscle phenotype MyoD gene transfer into skin fibroblasts has been attempted in an effort to diagnose genetic muscle diseases. Although the gene transduction efficiency of adenoviral gene delivery systems is higher than that of various other systems, the rate of myo-conversion is insignificant. Since high adenovirus doses are cytotoxic and exogenous MyoD expression is insufficient for skin fibroblasts to re-differentiate into muscle cells, we constructed the novel adeno-MyoD vector, Ad.CAGMyoD using the recombinant CAG promoter. Even at a lower multiplicities of infection most skin fibroblasts infected with Ad.CAGMyoD could convert into myotubes without vector-induced cytotoxicity. The converted cells expressed muscle-specific desmin and full-length dystrophin, both of which were detected by Western blotting. Genetic and immunohistochemical analyses using skin fibroblasts and our vector system are reliable and useful for the clinical diagnosis of genetic muscle diseases.

Selective binding of trisamine-modified phosphorothioate antisense DNA to target mRNA improves antisense activity and reduces toxicity.

Antisense activity in living cells has been thought to occur via a mechanism involving both DNA-mediated hybridization arrest of target mRNA and RNase H-mediated mRNA digestion. Therefore an ideal antisense agent should be permeable to the cell and possess capacities (1) to form a thermally stable duplex in vivo with its target, (2) to discriminate between mRNAs with different degrees of complementarity, and (3) to form antisense/RNA complexes that are susceptible to RNase H hydrolysis. A trisamine-modified deoxyuridine derivative of a novel phosphorothioate DNA 15-mer that meets all these criteria is described here. Compared with the unmodified phosphorothioate oligomer, the phosphorothioate derivative exhibits a higher antisense activity as well as reduced cytotoxicity in cells infected with HIV-1. Our data suggest that the melting temperature (T(m)) between antisense DNA and the target mRNA is not only one of the factors contributing to this derivative's improved antisense activity. Also important are an enhanced ability to discriminate between sequences and an increased susceptibility of the DNA/mRNA complex to RNase H hydrolysis. These results will be useful in designing more active, clinically useful antisense drugs.

Acute transverse myelitis after Japanese B encephalitis vaccination in a 4-year-old girl.

Fourteen days after Japanese B encephalitis (JBE) vaccination, a 4-year-old girl developed the full clinical manifestation of ATM within 24h. She showed acute ascending flaccid paraplegia with sensory disturbance, bladder dysfunction and meningeal sign. Cerebrospinal fluid examination revealed neutrophil pleocytosis and elevated protein level. Magnetic resonance imaging (MRI) showed diffuse swelling of the cervical and lumbar cord with low signal intensity on T1 and high signal intensity on T2-weighted imaging. These findings suggested that she had developed meningo-radiculomyelopathy. Since sequential MRI studies showed prompt reduction of the cord swelling, the high-dose methylprednisolone therapy employed seemed to have been effective for improvement of inflammation. Even with such potent drug treatment, she still has substantial flaccid diplegia and sphincter disorder 1 year later, and so we are convinced that the pathological change of the cord was as severe as in necrotizing myelopathy. Although the pathological process remains unknown, cellular autoimmune mechanism against the JBE vaccination is suspected.