Influences of Pinpoint Plantar Long-Wavelength Infrared Light Irradiation (Stress-Free Therapy) on Chorioretinal Hemodynamics, Atherosclerosis Factors, and Vascular Endothelial Growth Factor.

BACKGROUND: We previously reported that pinpoint plantar long-wavelength infrared light irradiation (stress-free therapy; SFT) is useful for alleviating insulin resistance and improving intracranial blood flow in patients with type 2 diabetes mellitus. This study was undertaken to evaluate the influences of SFT on chorioretinal hemodynamics (retinal artery and vein blood flows) as well as atherosclerosis-related factors (TG, LDL-C) and VEGF in patients with dyslipidemia. METHODS: Four patients with dyslipidemia received 15-minute irradiation with a stress-free apparatus (far-infrared wavelength, 30 mW). Using laser speckle flowgraphy, associations of chorioretinal blood flow with peripheral atherosclerosis-inducing factors/VEGF levels before and after irradiation were analyzed. RESULTS: Chorioretinal blood flow increased, while TG/LDL-C levels decreased, after irradiation. VEGF tended to rise in cases with pre-irradiation baseline levels at the lower limit but tended to decrease in cases in which baseline levels had exceeded the normal range. CONCLUSION: SFT was suggested to enhance chorioretinal circulation and to normalize VEGF, thereby possibly contributing to amelioration of atherosclerosis-inducing factors. Abnormalities in chorioretinal hemodynamics are known to be highly involved in the pathophysiology of diabetic retinopathy and age-related macular degeneration, and anti-VEGF antibody has been used for treating these conditions. The necessity of risk management, involving chorioretinal blood flow, has been pointed out when dealing with central retinal vein occlusion, diabetes mellitus, ischemic cerebral/cardiac disease, dementia and so on. SFT is therefore a potential complementary medical strategy which can be expected to contribute to normalization of chorioretinal blood flow and atherosclerosis-inducing factors/VEGF levels, and thereby to the prevention of lifestyle-related chronic diseases.

Effect of Stress-Free Therapy on immune system: Induction of Interleukin 10 expression in lymphocytes through activation of CD19(+) CD24(hi) CD38(hi) regulatory B Cells.

BACKGROUND AND AIMS: Mild thermal treatment with "Pinpoint Plantar Long-wavelength Infrared Light Irradiation (PP-LILI)" named as Stress-Free Therapy(®) increases peripheral-deep body temperature and blood flow, and improves multiple disorders including hyperpiesia, type II diabetes and cardiovascular patients. Immunomodulatory effects of PP-LILI were investigated. MATERIALS AND METHODS: Seven healthy individuals and 4 people with underlying medical condition (UMC) participated in this study. Participants were given PP-LILI stimuli twice a week over 3 weeks and followed with placebo stimuli over 3 weeks. This set of sessions was repeated 3 times. For analyses, fresh peripheral mononuclear cells from participants were stained with fluorescencedye conjugated monoclonal antibodies and changes in populational compositions and IL-10 expression levels were observed by flow cytometry. RESULTS: Distinct expression of IL-10 in lymphocytes was induced by PP-LILI from the second session in the healthy individuals. This induction was terminated during the following placebo sessions. PP-LILI induced activation of CD19(+) CD24(hi) CD38(hi) regulatory B cells in every session prior to induce the IL-10 in major lymphocytes. Activated regulatory B cells in the individuals with UMC decreased as same levels of healthy individuals after second PP-LILI session and re-activated with the stimuli. Significant population changes in neither regulatory T cells nor proinflammatory IL-17A expressing CD4(+) T cells were observed. CONCLUSIONS: PP-LILI is a potent immunomodulatory inducer that activates regulatory B cells and consequent IL-10 expression in lymphocytes. Moreover, its stimulatory intervals down-regulate the higher activation of regulatory B cells and lymphocyte's IL-10 expression occurred by UMC to the healthy people's level.

Preliminary results of highly localized plantar irradiation with low incident levels of mid-infrared energy which contributes to the prevention of dementia associated with underlying diabetes mellitus.

BACKGROUND AND AIMS: The incidence of vascular dementia (VD) and Alzheimer's disease (AD) has recently increased and the prevention of progression of these diseases is very difficult. RESULTS: The application of pinpoint plantar long-wavelength infrared light irradiation (PP-LILI) to a patient's sole, at the point where the line drawn between the first and second metatarsal heads intersects with the vertical line from the medial malleolus, was effective in increasing blood flow to the facial artery, elevating high-density lipoprotein cholesterol (HDL-C) levels, and reducing insulin resistance. CONCLUSIONS: We found that these effects of PP-LILI might be helpful for preventing VD and AD, conditions that are becoming a social problem in an aging Japanese society.

[Corrigendum] Induction of antigen-specific cytotoxic T lymphocytes by fusion cells generated from allogeneic plasmacytoid dendritic and tumor cells.

Some errors in Fig. 3B and Fig. 6C have been identified. The errors do not change the conclusion of the paper. The conclusion is supported by other figures in the paper, as well as results described in the text. The corrected Fig. 3B and Fig. 6C are shown below. [the original article was published in the International Journal of Oncology 45: 470-478, 2014 DOI: 10.3892/ijo.2014.2433]

Immunogenic modulation of cholangiocarcinoma cells by chemoimmunotherapy.

BACKGROUND/AIM: Chemoimmunotherapy has been used to treat intrahepatic cholangiocarcinoma (ICC). However, little is known about the phenomena underlying the immunomodulation of ICC cells elicited by chemoimmunotherapy. MATERIALS AND METHODS: Primary ICC cells from a patient with ICC who received gemcitabine followed by 5-fluorouracil (5-FU), both combined with dendritic cells pulsed with Wilms' tumor 1 (WT1) peptides were cultured. ICC cells were treated with gemcitabine, 5-FU or interferon (IFN)-γ in vitro. The phenotype of the ICC cells was examined by flow cytometry and quantitative reverse transcription polymerase chain reaction. RESULTS: Stimulation of the ICC cells with gemcitabine resulted in up-regulation of WT1 mRNA, programmed death receptor ligand-1 (PDL1) and calreticulin. Gemcitabine, 5-FU and IFN-γ induced up-regulation of mucin-1. Moreover, human leukocyte antigen (HLA)-ABC, HLA-DR and PDL1 were extremely up-regulated by IFN-γ. CONCLUSION: Chemoimmunomodulating agents alter the immunogenicity of ICC cells, resulting in complex clinical efficacy results.

Hybrid micro-particles as a magnetically-guidable decontaminant for cesium-eluted ash slurry.

Decontamination of the radioactive cesium that is widely dispersed owing to a nuclear power station accident and concentrated in fly ash requires an effective elimination system. Radioactive fly ash contains large amounts of water-soluble cesium that can cause severe secondary contamination and represents a serious health risk, yet its complete removal is complicated and difficult. Here it is shown that a new fine-powder formulation can be magnetically guided to eliminate cesium after being mixed with the ash slurry. This formulation, termed MagCE, consists of a ferromagnetic porous structure and alkaline- and salt-resistant nickel ferrocyanide. It has potent cesium-adsorption- and magnetic-separation-properties. Because of its resistance against physical and chemical attack such as with ash particles, as well as with the high pH and salt concentration of the ash slurry, MagCE simplifies the decontamination process without the need of the continued presence of the hazardous water-soluble cesium in the treated ash.

Induction of antigen-specific cytotoxic T lymphocytes by fusion cells generated from allogeneic plasmacytoid dendritic and tumor cells.

Previous work has demonstrated that fusion cells generated from autologous monocyte-derived dendritic cells (MoDCs) and whole tumor cells induce efficient antigen-specific cytotoxic T lymphocytes. A major limitation to the use of this strategy is the availability of adequate amounts of autologous tumor cells. Moreover, MoDCs from cancer patients are often defective in their antigen-processing and presentation machinery. In this study, two types of allogeneic cells, a leukemia plasmacytoid dendritic cell (pDC) line (PMDC05) and pancreatic cancer cell lines (PANC-1 or MIA PaCa-2), were fused instead of autologous MoDCs and tumor cells. We created four types of pDC/tumor fusion cells by alternating fusion partners and treating with lipopolysaccharide (LPS): i) PMDC05 fused with PANC-1 (pDC/PANC-1), ii) PMDC05 fused with MIA PaCa-2 (pDC/MIA PaCa-2), iii) LPS-stimulated pDC/PANC-1 (LPS-pDC/PANC-1) and iv) LPS-stimulated pDC/MIA PaCa-2 (LPS-pDC/MIA PaCa-2) and examined their antitumor immune responses. The LPS-pDC/tumor cell fusions were the most active, as demonstrated by their: i) upregulated expression of HLA-DR and CD86 on a per-fusion-cell basis, ii) increased production of IL-12p70, iii) generation of a higher percentage of IFN-γ-producing CD4⁺ and CD8⁺ T cells and iv) augmented induction of MUC1-specific CD8⁺ T cells that lyse target tumor cells. This study provides the first evidence for an in vitro induction of antigen-specific cytotoxic T lymphocytes by LPS-stimulated fusion cells generated from leukemia plasmacytoid DCs and tumor cells and suggests that this strategy has potential applicability to the field of adoptive immunotherapy.

Effect of Stress-free Therapy on Cerebral Blood Flow: Comparisons among patients with metabolic cardiovascular disease, healthy subjects and placebo-treated subjects.

BACKGROUND AND AIMS: We have developed a Stress-free Therapy(®) device wherein "Pinpoint Plantar Long-wavelength Infrared Light Irradiation (PP-LILI)" increases peripheral-deep body temperature and blood flow volume and stabilizes blood pressure as well as significantly reduces stress hormones such as adrenocorticotrophic hormone and cortisol without using drugs. Moreover, we have found this therapy to significantly improve blood glucose and insulin resistance in patients with type 2 diabetes. Based on this background of clinical efficacy, we validated changes in cerebral blood flow in patients with metabolic cardiovascular disease and examined the efficacy of Stress-free Therapy(®) on cerebral blood flow as compared to that in healthy control subjects and placebo-treated patients. RESULTS: The change in cerebral blood flow volume during 15-minute PP-LILI was 5.1 ± 1.8 mL/min in patients with metabolic cardiovascular disease, showing a significant increase (P<0.05) of 3.1 mL/min as compared with the mean blood flow value after resting for 15 minutes. CONCLUSIONS: Our results suggested Stress-free Therapy(®) to significantly increase cerebral blood flow, possibly leading to the prevention of metabolic cardiovascular disease.

Strategies to improve the immunogenicity of anticancer vaccines based on dendritic cell/malignant cell fusions.

The rationale for fusing dendritic cells (DCs) with whole tumor cells to generate anticancer vaccines resides in the fact that the former operate as potent antigen-presenting cells, whereas the latter express a constellation of tumor-associated antigens (TAAs). Although the administration of DC/malignant cell fusions to cancer patients is safe and this immunotherapeutic intervention triggers efficient tumor-specific T-cell responses in vitro, a limited number of objective clinical responses to DC/cancer cell fusions has been reported thus far. This review discusses novel approaches to improve the immunogenicity of DC/malignant cell fusions as anticancer vaccines.

Preliminary results of pinpoint plantar long-wavelength infrared light irradiation on blood glucose, insulin and stress hormones in patients with type 2 diabetes mellitus.

BACKGROUND AND AIMS: This study was aimed at the development of a novel noninvasive treatment system, "pinpoint plantar long-wavelength infrared light irradiation (PP-LILI)", which may be able to relieve mental stress and normalize blood glucose level via the reduction of stress hormones in type 2 (non-insulin-dependent) diabetes mellitus (DM) patients. MATERIALS (SUBJECTS) AND METHODS: Based on this hypothesis, the present study was undertaken to examine effects of PP-LILI on stress hormones (ACTH and cortisol), blood glucose, HbA1c, and insulin levels in 10 patients with type 2 DM. Each patient received PP-LILI of the foot for 15 minutes once weekly using a stress free apparatus (infrared wavelength, 9,000-12,000 nm/power 30 mW). RESULTS: In response to this therapy, ACTH (P<0.01) and cortisol (P<0.05) levels decreased significantly. Fasting blood glucose (P<0.05) and insulin (P<0.05) levels also decreased significantly along with a tendency for HbA1c to decrease. CONCLUSIONS: The present data raise the possibility that PP-LILI can normalize blood glucose levels by reducing stress hormones such as cortisol, which aggravate DM, and by improving insulin sensitivity, thereby contributing to prevention and treatment of DM.

Effect of pinpoint plantar long-wavelength infrared light irradiation on subcutaneous temperature and stress markers.

BACKGROUND AND AIMS: The current investigation was aimed at the development of a novel non-invasive treatment system, "pinpoint plantar long-wavelength infrared light irradiation (PP-LILI)", which may be able to relieve mental stress and reduce stress-related hormones. Materials (Subjects) and methods: We compared the subcutaneous temperature, blood pressure, the degree of secretion of stress hormones before and after pinpoint irradiations (wavelength: 8-11 µm; output: 30mW). The study enrolled 15 subjects (Japanese healthy adults; 8 males, 7 females; average age 47.8 ± 14.6 years). Two parts of the planter region were irradiated for 15 min respectively. The stress markers such as ACTH, salivary amylase and cortisol were measured. As well, core body temperature and blood pressure were analyzed before and after the irradiation. RESULTS: A series of experiments revealed increased body temperature, decreased levels of blood pressure and stress markers described above after the irradiation. CONCLUSIONS: These results clearly suggest that the PP-LILI system will be quite useful for relieving stress and improvement of homeostatic functions in the body.

Improved immunogenicity of fusions between ethanol-treated cancer cells and dendritic cells exposed to dual TLR stimulation.

Tumor-derived transforming growth factor ß1 (TGFß1) generally abrogates the immunogenicity of dendritic cells (DCs) fused to whole cancer cells. We have recently revealed that ethanol-treated neoplastic cells fused to DCs exposed to 2 Toll-like receptor agonists efficiently induce cytotoxic T lymphocytes via TGFß1 blockade and the production of interleukin-12.

The combination of TLR2 and TLR4 agonists promotes the immunogenicity of dendritic cell/cancer cell fusions.

The induction of antitumor immune responses by dendritic cell (DC)/tumor cell fusions can be modulated by their activation status. Our recent work reveals that the combination of Toll-like receptor 2 (TLR2) and TLR4 agonists promotes the immunogenicity of DC/tumor cell fusions, allowing them to overcome the immunosuppressive effects of transforming growth factor ß1.

Fusions between dendritic cells and whole tumor cells as anticancer vaccines.

Various strategies have been developed to deliver tumor-associated antigens (TAAs) to dendritic cells (DCs). Among these, the fusion of DCs and whole cancer cells can process a broad array of TAAs, including hitherto unidentified molecules, and present them in complex with MHC Class I and II molecules and in the context of co-stimulatory signals. DC-cancer cell fusions have been shown to stimulate potent antitumor immune responses in animal models. In early clinical trials, however, the antitumor effects of DC-cancer cell fusions are not as vigorous as in preclinical settings. This mini-review summarizes recent advances in anticancer vaccines based on DC-cancer cell fusions.

Augmentation of antitumor immunity by fusions of ethanol-treated tumor cells and dendritic cells stimulated via dual TLRs through TGF-ß1 blockade and IL-12p70 production.

The therapeutic efficacy of fusion cell (FC)-based cancer vaccine generated with whole tumor cells and dendritic cells (DCs) requires the improved immunogenicity of both cells. Treatment of whole tumor cells with ethanol resulted in blockade of immune-suppressive soluble factors such as transforming growth factor (TGF)-ß1, vascular endothelial growth factor, and IL-10 without decreased expression of major histocompatibility complex (MHC) class I and the MUC1 tumor-associated antigen. Moreover, the ethanol-treated tumor cells expressed "eat-me" signals such as calreticulin (CRT) on the cell surface and released immunostimulatory factors such as heat shock protein (HSP)90α and high-mobility group box 1 (HMGB1). A dual stimulation of protein-bound polysaccharides isolated from Coriolus versicolor (TLR2 agonist) and penicillin-inactivated Streptococcus pyogenes (TLR4 agonist) led human monocyte-derived DCs to produce HSP90α and multiple cytokines such as IL-12p70 and IL-10. Interestingly, incorporating ethanol-treated tumor cells and TLRs-stimulated DCs during the fusion process promoted fusion efficiency and up-regulated MHC class II molecules on a per fusion basis. Moreover, fusions of ethanol-treated tumor cells and dual TLRs-stimulated DCs (E-tumor/FCs) inhibited the production of multiple immune-suppressive soluble factors including TGF-ß1 and up-regulated the production of IL-12p70 and HSP90α. Most importantly, E-tumor/FCs activated T cells capable of producing high levels of IFN-γ, resulting in augmented MUC1-specific CTL induction. Collectively, our results illustrate the synergy between ethanol-treated whole tumor cells and dual TLRs-stimulated DCs in inducing augmented CTL responses in vitro by FC preparations. The alternative system is simple and may provide a platform for adoptive immunotherapy.

Combined TLR2/4-activated dendritic/tumor cell fusions induce augmented cytotoxic T lymphocytes.

Induction of antitumor immunity by dendritic cell (DC)-tumor fusion cells (DC/tumor) can be modulated by their activation status. In this study, to address optimal status of DC/tumor to induce efficient antigen-specific cytotoxic T lymphocytes (CTLs), we have created various types of DC/tumor: 1) un-activated DC/tumor; 2) penicillin-killed Streptococcus pyogenes (OK-432; TLR4 agonist)-activated DC/tumor; 3) protein-bound polysaccharides isolated from Coriolus versicolor (PSK; TLR2 agonist)-activated DC/tumor; and 4) Combined OK-432- and PSK-activated DC/tumor. Moreover, we assessed the effects of TGF-ß1 derived from DC/tumor on the induction of MUC1-specific CTLs. Combined TLR2- and TLR4-activated DC/tumor overcame immune-suppressive effect of TGF-ß1 in comparison to those single activated or un-activated DC/tumor as demonstrated by: 1) up-regulation of MHC class II and CD86 expression on DC/tumor; 2) increased fusion efficiency; 3) increased production of fusions derived IL-12p70; 4) activation of CD4(+) and CD8(+) T cells that produce high levels of IFN-γ; 5) augmented induction of CTL activity specific for MUC1; and 6) superior efficacy in inhibiting CD4(+)CD25(+)Foxp3(+) T cell generation. However, DC/tumor-derived TGF-ß1 reduced the efficacy of DC/tumor vaccine in vitro. Incorporating combined TLRs-activation and TGF-ß1-blockade of DC/tumor may enhance the effectiveness of DC/tumor-based cancer vaccines and have the potential applicability to the field of adoptive immunotherapy.

Synthesis of lipidic magnetic nanoparticles for nucleic acid delivery.

Lipidic magnetic nanoparticles are a rapid and reliable magnet-guided nucleic acid delivery system, which we have named as "LipoMag" (Namiki et al., Nat. Nanotechnol 4:598-606, 2009). LipoMag is composed of a cationic lipid shell, an oleic acid binder, and an iron oxide nanocrystal core. Through the electrostatic force, positively charged LipoMag and negatively charged nucleic acid form complexes, termed "mag-lipoplex." Under a magnetic field, LipoMag bearing nucleic acid can be controllably accumulated in target cells. Here we describe the procedure for the preparation of LipoMag which can be used for the transfection of nucleic acids, such as plasmid DNA and small interfering RNA.

Immunotherapy for colorectal cancer.

The incidence of colorectal cancer (CRC) is on the rise, and the prognosis for patients with recurrent or metastatic disease is extremely poor. Although chemotherapy and radiation therapy can improve survival rates, it is imperative to integrate alternative strategies such as immunotherapy to improve outcomes for patients with advanced CRC. In this review, we will discuss the effect of immunotherapy for inducing cytotoxic T lymphocytes and the major immunotherapeutic approaches for CRC that are currently in clinical trials, including peptide vaccines, dendritic cell-based cancer vaccines, whole tumor cell vaccines, viral vector-based cancer vaccines, adoptive cell transfer therapy, antibody-based cancer immunotherapy, and cytokine therapy. The possibility of combination therapies will also be discussed along with the challenges presented by tumor escape mechanisms.

Several factors including ITPA polymorphism influence ribavirin-induced anemia in chronic hepatitis C.

AIM: To construct formulae for predicting the likelihood of ribavirin-induced anemia in pegylated interferon α plus ribavirin for chronic hepatitis C. METHODS: Five hundred and sixty-one Japanese patients with hepatitis C virus genotype 1b who had received combination treatment were enrolled and assigned randomly to the derivation and confirmatory groups. Single nucleotide polymorphisms at or nearby ITPA were genotyped by real-time detection polymerase chain reaction. Factors influencing significant anemia (hemoglobin concentration < 10.0 g/dL at week 4 of treatment) and significant hemoglobin decline (declining concentrations > 3.0 g/dL at week 4) were analyzed using multiple regression analyses. Prediction formulae were constructed by significantly independent factors. RESULTS: Multivariate analysis for the derivation group identified four independent factors associated with significant hemoglobin decline: hemoglobin decline at week 2 [P = 3.29 × 10(-17), odds ratio (OR) = 7.54 (g/dL)], estimated glomerular filtration rate [P = 2.16 × 10(-4), OR = 0.962 (mL/min/1.73 m(2))], rs1127354 (P = 5.75 × 10(-4), OR = 10.94) and baseline hemoglobin [P = 7.86 × 10(-4), OR = 1.50 (g/dL)]. Using the model constructed by these factors, positive and negative predictive values and predictive accuracy were 79.8%, 88.8% and 86.2%, respectively. For the confirmatory group, they were 83.3%, 91.0% and 88.3%. These factors were closely correlated with significant anemia. However, the model could not be constructed, because no patients with rs1127354 minor genotype CA/AA had significant anemia. CONCLUSION: Reliable formulae for predicting the likelihood of ribavirin-induced anemia were constructed. Such modeling may be useful in developing individual tailoring and optimization of ribavirin dosage.

Contribution of ribavirin transporter gene polymorphism to treatment response in peginterferon plus ribavirin therapy for HCV genotype 1b patients.

BACKGROUND: Standard-dose ribavirin is crucial for the standard-of-care treatment of chronic hepatitis C virus (HCV) infection. Equilibrative nucleoside transporter 1 (ENT1), encoded by SLC29A1 gene, is the main transporter that imports ribavirin into human hepatocytes. AIMS: To determine whether single nucleotide polymorphisms (SNPs) at the SLC29A1 gene could influence the probability of treatment response compared with other baseline and host genetic factors. METHODS: A total of 526 East Asian patients monoinfected with HCV genotype 1b who had received pegylated interferon alpha plus ribavirin therapy were enrolled in this study. They were assigned randomly to the derivation and confirmatory groups. SNPs related to the IL28B, ITPA and SLC29A1 genes were genotyped using real-time detection polymerase chain reaction. Factors associated with sustained virological response (SVR) were analysed using multiple logistic regression analysis. RESULTS: Multivariate analysis for the derivation group identified six baseline variables significantly and independently associated with SVR: age [P = 0.023, odds ratio (OR) = 0.97], gender (P = 0.0047, OR = 2.25), platelet count (P = 0.00017, OR = 1.11), viral load (P = 0.00026, OR = 0.54), IL28B SNP rs12979860 (P = 1.09 × 10(-7) , OR = 8.68) and SLC29A1 SNP rs6932345 (P = 0.030, OR = 1.85). Using the model constructed by these independent variables, positive and negative predictive values and predictive accuracy were 73.3, 70.1 and 71.9% respectively. For the confirmatory group, they were 71.4, 84.6 and 75.3% respectively. The SLC29A1 and IL28B SNPs were also significantly associated with rapid virological response. CONCLUSIONS: The SNP at the major ribavirin transporter ENT1 gene SLC29A1 was one of significantly independent factors influencing treatment response, although the impact on the prediction was small.