Antitumour and Apoptotic Effects of a Plant Extract Mixture Containing Rhus verniciflua and Other Herbs in Human Leukaemia Cells.

The apoptotic effects of a novel antitumour agent (Rv-PEM01) prepared from 6 kinds of herbs, including Rhus verniciflua were investigated using flow cytometry and western blot analysis. Rv-PEM01 induced apoptosis but not necrosis in MOLT-3, KG-1, and K562 human leukaemia cell lines. Further, Rv-PEM01-treated cells showed significantly upregulated expression of caspase-3 and 9 and cleaved caspase-3 and 9 compared to the control cells. Taken together, the results suggest that Rv-PEM01 induced apoptosis via the mitochondrial-mediated pathway, and is a potential natural anticancer agent and/or a functional food material.

Mao-to Prolongs the Survival of and Reduces TNF-alpha Expression in Mice with Viral Myocarditis.

Goal of this study was to evaluate effects of Mao-to on development of myocarditis induced by encephalomyocarditis (EMC) virus in mice. Mice were randomly divided into five groups. Group N included uninfected controls (n = 18), while group A, B and C underwent intraperitoneal injection of EMC virus. Group A was administered oral saline from day 0 to day 4. Group B was administered oral Mao-to (500 mg(-1) kg(-1) day(-1)) from day 0 to day 4. Group C was administered Mao-to from day 2 to day 6. Group D was administered Mao-to from day 5 to day 10. Treated mice were followed for survival rates during 2 weeks after infection. Body weight (BW) and organ weights including heart (HW), lungs, thymus and spleen were examined on days 4, 6 and 14. Survival rate of group C (36.4%) was significantly improved compared with group A, B or D (0% of each, P < 0.05). HW and HW/BW ratio in group C was significantly (P < 0.05) lower than those in group A, B or D. Viral titers of hearts were significantly different among groups A, B and C. Cardiac expression in tumor necrosis factor-alpha (TNF-alpha) was significantly reduced in group C in comparison with group A, B or D on day 6 by immunohistochemical study. Administration of Mao-to starting on day 2 improves mortality resulting from viral myocarditis in mice with reduced expression of cardiac TNF-alpha. These findings suggest that timing of Mao-to is crucial for preventing cardiac damage in mice with viral myocarditis.

Aspects of sino-Japan complementary and alternative medicine and development on the traditional uighur medicine.

Two consecutive conferences on 'Sino-Japan Complementary and Alternative Medicine and Development on the Traditional Uighur Medicine' were held in Xinjiang Medical University on July 3 and Kanazawa Medical University on October 6, 2007. The Vice president Halmurat Upur presided over the meeting and gave congratulatory address on holding of the conference. In order to understand mutually and discuss the possibility of the Uighur Medicine as CAM and the situation of medicine in the global sense, specialist scholars of Traditional Uighur Medicine and postgraduates attended this conference. In the meeting of the CAM, the achievements on the research of Traditional Uighur Medicine were exchanged and warmly discussed. Presentations were made in the consecutive conference.

Do Japanese style acupuncture and moxibustion reduce symptoms of the common cold?

We summarize the results from a series of investigations of Japanese style acupuncture and moxibustion therapies on symptoms of the common cold that have been conducted (FTLE 1999-03, supported by the Foundation for Training and Licensure Examination in Anma- Massage- Acupressure, Acupuncture and Moxibustion). We also discuss the various interventions and concerns that we faced during these investigations. The subjects were students and teachers. The pilot study (FTLE1999) of a two arm (real and non-treatment control) RCT at a Japanese acupuncture school showed that manual acupuncture to a specific needling point at the throat clearly reduced symptoms of the common cold. The first multi-center (five centers) RCT (FTLE 2000) revealed a significant reduction in cold symptoms, by general linear model analysis (between groups, P = 0.024). To reduce the technical variation, we employed indirect moxibustion to the neck points as a uniform intervention in the next project (FTLE 2001) without statistically significant results. Then we elongated the periods of treatment from 2 to a maximum of 12 weeks (FTLE 2002) with different interventions accompanied by 4 weeks follow-up. The results were still not statistically significant. As the final project, we tried to develop a new experimental design for individualized intervention by conducting n-of-1 trials using elderly subjects in a health care center but without detecting a clear effect. In conclusion, the safety of Japanese acupuncture or moxibustion was sufficiently demonstrated; however, a series of clinical trials could not offer convincing evidence to recommend the use of Japanese style acupuncture or moxibustion for preventing the common cold. Further studies are required as the present trials had several limitations.

Brain atrophy in a murine model of chronic fatigue syndrome and beneficial effect of Hochu-ekki-to (TJ-41).

Brain-derived neurotrophic factor (BDNF) is associated with the main symptoms of chronic fatigue syndrome (CFS) and neuron apoptosis. Nevertheless, no study has been performed directly to explore the relationship between CFS, BDNF and neuron apoptosis. We induced a CFS model by six injections of killed Brucella abortus antigen in BALB/c mice and treated them with Hochu-ekki-to (TJ-41). Daily running activity, body weight (BW), ratio of cerebral weight to BW (CW/BW) and expression levels of BDNF and Bcl-2 mRNA in the hippocampus were determined. The daily activity and CW/BW decreased significantly in the CFS model. BDNF and Bcl-2 mRNA expression levels in the hippocampus were suppressed in the CFS model and TJ-41 treated mice, while no significant difference was found between them. We improved a murine model to investigate the relationship between CFS and brain dysfunction. In this model, reduced daily activity might have been associated with decreased hippocampal BDNF mRNA expression, hippocampal apoptosis and brain atrophy. TJ-41 increased the daily running activity of the model, which was independent of brain recovery.

Characterization of a novel vanadium-binding protein (VBP-129) from blood plasma of the vanadium-rich ascidian Ascidia sydneiensis samea.

The ascidians, the so-called sea squirts, accumulate high levels of vanadium, a transition metal. Since Henze first observed this physiologically unusual phenomenon about one hundred years ago, it has attracted interdisciplinary attention from chemists, physiologists, and biochemists. The maximum concentration of vanadium in ascidians can reach 350 mM, and most of the vanadium ions are stored in the +3 oxidation state in the vacuoles of vanadium-accumulating blood cells known as vanadocytes. Many proteins involved in the accumulation and reduction of vanadium in the vanadocytes, blood plasma, and digestive tract have been identified. However, the process by which vanadium is taken in prior to its accumulation in vanadocytes has not been elucidated. In the present study, a novel vanadium-binding protein, designated VBP-129, was identified from blood plasma of the vanadium-rich ascidian Ascidia sydneiensis samea. Although VBP-129 mRNA was transcribed in all A. sydneiensis samea tissues examined, the VBP-129 protein was exclusively localized in blood plasma and muscle cells of this ascidian. It bound not only to VO(2+) but also to Fe(3+), Co(2+), Cu(2+), and Zn(2+); on the other hand, a truncated form of VBP-129, designated VBP-88, bound only to Co(2+), Cu(2+) and Zn(2+). In a pull-down assay, an interaction between VanabinP and VBP-129 occurred both in the presence and the absence of VO(2+). These results suggest that VBP-129 and VanabinP function cooperatively as metallochaperones in blood plasma.

Acupuncture regulates leukocyte subpopulations in human peripheral blood.

Acupuncture has recently been attracting more and more people throughout the world as an alternative treatment, however little is known about its physiological activities (i.e. immune system). We examined acupuncture both quantitatively and qualitatively by measuring CD-positive cell counts and cytokine expression levels in the blood, to determine the activity of T cells, B cells, macrophages and natural killer (NK) cells. Fifteen milliliters of peripheral blood obtained from 17 healthy volunteers aged 21-51 years, were analyzed using flow cytometry before and after acupuncture treatment. There was a statistically significant increase in the number of CD2(+), CD4(+), CD8(+), CD11b(+), CD16(+), CD19(+), CD56(+) cells as well as IL-4, IL-1beta and IFN-gamma levels in the cells after acupuncture stimulation of meridian points. These observations indicate that acupuncture may regulate the immune system and promote the activities of humoral and cellular immunity as well as NK cell activity. In this article, we discussed how acupuncture regulated leukocyte numbers and functions since they are considered to be potential indicators for evaluating complementary and alternative medicine.

Inhibition of ganciclovir-resistant human cytomegalovirus replication by Kampo (Japanese herbal medicine).

We examined the effect of Kampo on the replication of ganciclovir (GCV)-resistant human cytomegalovirus (HCMV) in the human embryonic fibroblast cell line MRC-5. Treatment of HCMV-infected cells with Sho-seiryu-to (SST; Xiao-Qing-Long-Tang in Chinese) resulted in the inhibition of viral replication without affecting the cell growth. SST treatment decreased the synthesis of viral DNA, but had no virucidal effect on cell-free HCMV. However, the inhibitory effect of SST on HCMV replication was ablated by anti-interferon-beta (IFN-beta) antibody suggesting that SST inhibits the replication of GCV-resistant HCMV through the induction of IFN-beta. These results suggest that SST is a novel compund with potential as an anti-HCMV.

Beneficial effect of brewers' yeast extract on daily activity in a murine model of chronic fatigue syndrome.

The aim of this study was to assess the effect of Brewers' yeast extract (BYE) on daily activity in a mouse model of chronic fatigue syndrome (CFS). CFS was induced by repeated injection of Brucella abortus (BA) antigen every 2 weeks. BYE was orally administered to mice in a dose of 2 g per kg per day for 2 weeks before injecting BA and for 4 weeks thereafter. We evaluated daily running activity in mice receiving BYE as compared with that in untreated mice. Weekly variation of body weight (BW) and survival in both groups was monitored during the observation period. Spleen weight (SW), SW/BW ratio, percent splenic follicular area and expression levels of interferon-gamma (IFN-gamma) and interleukin-10 (IL-10) mRNA in spleen were determined in both groups at the time of sacrifice. The daily activity during 2 weeks after the second BA injection was significantly higher in the treated group than in the control. There was no difference in BW between both groups through the experimental course. Two mice in the control died 2 and 7 days after the second injection, whereas no mice in the treated group died. Significantly decreased SW and SW/BW ratio were observed in the treated mice together with elevation of splenic follicular area. There were suppressed IFN-gamma and IL-10 mRNA levels in spleens from the treated mice. Our results suggest that BYE might have a protective effect on the marked reduction in activity following repeated BA injection via normalization of host immune responses.

VanabinP, a novel vanadium-binding protein in the blood plasma of an ascidian, Ascidia sydneiensis samea.

Some ascidians accumulate high levels of the transition metal vanadium in their blood cells. The process of vanadium accumulation has not yet been elucidated. In this report, we describe the isolation and cDNA cloning of a novel vanadium-binding protein, designated as VanabinP, from the blood plasma of the vanadium-rich ascidian, Ascidia sydneiensis samea. The predicted amino acid sequence of VanabinP was highly conserved and similar to those of other Vanabins. The N-terminus of the mature form of VanabinP was rich in basic amino acid residues. VanabinP cDNA was originally isolated from blood cells, as were the other four Vanabins. However, Western blot analysis revealed that the VanabinP protein was localized to the blood plasma and was not detectable in blood cells. RT-PCR analysis and in situ hybridization indicated that the VanabinP gene was transcribed in some cell types localized to peripheral connective tissues of the alimentary canal, muscle, blood cells, and a portion of the branchial sac. Recombinant VanabinP bound a maximum of 13 vanadium(IV) ions per molecule with a Kd of 2.8 x 10(-5) M. These results suggest that VanabinP is produced in several types of cell, including blood cells, and is immediately secreted into the blood plasma where it functions as a vanadium(IV) carrier.

In Vitro Anti-Cytomegalovirus Activity of Kampo (Japanese Herbal) Medicine.

We examined the effect of three types of Kampo medicines on human cytomegalovirus (CMV) replication in the human embryonic fibroblast cell line, MRC-5. Treatment of cells with at least 0.01 mug/ml of Kampo medicines inhibited the cytopathic effects of CMV-infected MRC-5 cells. Moreover, Kampo medicine decreased the replication of CMV without affecting the inhibition of host cells, with a concomitant decrease in CMV DNA levels. However, Kampo medicine demonstrated no virocidal effect on cell-free CMV. Furthermore, western blotting analysis demonstrated that the Kampo medicine decreased the amount of 65 kDa late antigen expression in the infected cells. These results suggest that Kampo medicine may be sufficient to inhibit viral DNA replication and late protein synthesis, resulting in anti-CMV effects. Therefore, these three Kampo medicines have the potential of being a source of new powerful anti-CMV compounds.

Preventive and curative effects of acupuncture on the common cold: a multicentre randomized controlled trial in Japan.

OBJECTIVE: To determine the preventive and curative effects of manual acupuncture on the symptoms of the common cold. METHOD: Students and staff in five Japanese acupuncture schools (n=326) were randomly allocated to acupuncture and no-treatment control groups. A specific needling point (Y point) on the neck was used bilaterally. Fine acupuncture needles were gently manipulated for 15 s, evoking de qi sensation. Acupuncture treatments were performed four times during the 2-week experimental period with a 2-week follow-up period. A common cold diary was scored daily for 4 weeks, and a common cold questionnaire was scored before each acupuncture treatment and twice at weekly intervals. A reliability test for the questionnaire was performed on the last day of recording. RESULTS: Five of the 326 subjects who were recruited dropped out. The diary score in the acupuncture group tended to decrease after treatment, but the difference between groups was not significant (Kaplan-Meier survival analysis, log rank test P=0.53, Cox regression analysis, P>0.05). Statistically significantly fewer symptoms were reported in the questionnaire by the acupuncture group than control group (P=0.024, general linear model, repeated measure). Significant inter-centre (P<0.001, general linear model) and sex (P=0.027, general linear model) differences were also detected. Reliability tests indicated that the questionnaire with 15 items was sufficiently reliable. No severe adverse event was reported. CONCLUSION: This is the first report of a multi-centre randomized controlled trial of acupuncture for symptoms of the common cold. A significantly positive effect of acupuncture was demonstrated in the summed questionnaire data, although a highly significant inter-centre difference was observed. Needling on the neck using the Japanese fine needle manipulating technique was shown to be effective and safe. The use of acupuncture for symptoms of the common cold symptoms should be considered, although further evidence from placebo controlled RCTs is required.

Expressed sequence tag analysis of vanadocytes in a vanadium-rich ascidian, Ascidia sydneiensis samea.

Some species in the family Ascidiidae accumulate vanadium at concentrations in excess of 350 mM, which corresponds to about 10(7) times higher than that in seawater. In these species signet ring cells, with a single huge vacuole in which vanadium ion is contained, function as vanadium-accumulating cells, vanadocytes. To investigate the mechanism underlying this phenomenon, we performed an expressed sequence tag (EST) analysis of a complementary DNA library from vanadocytes of a vanadium-rich ascidian, Ascidia sydneiensis samea. We determined the nucleotide sequences of 1000 ESTs and performed a BLAST analysis against the SwissProt database. We found 93 clones of metal-related gene homologues, including the ferritin heavy subunit, hemocyanin, and metallothionein. Two ESTs, in particular, exhibited significant similarity to vanabins that have been extracted from A. sydneiensis samea blood cells as low molecular weight vanadium-binding proteins. We have named the genes encoding these ESTs vanabin3 and vanabin4. Immobilized metal ion affinity chromatography revealed that these novel vanabin homologues bind vanadium(IV) ions.

Novel vanadium-binding proteins (vanabins) identified in cDNA libraries and the genome of the ascidian Ciona intestinalis.

Ascidians, especially those belonging to the suborder Phlebobranchia, can accumulate high levels of vanadium. Vanadium-binding proteins (vanabins) were first isolated from a vanadium-accumulating ascidian, Ascidia sydneiensis samea, and then the vanabins were cloned, their expression was studied, and metal-binding assays were conducted. In order to unravel the mechanism of vanadium accumulation, we searched for vanabin-like genes in other animals, including other ascidians. A database search revealed five groups of cDNAs that encoded vanabin-like proteins in another ascidian, Ciona intestinalis. The genes encoding C. intestinalis vanabins, CiVanabin1 to CiVanabin5, were clustered in an 8.4-kb genomic region. The direction of transcription of each gene was identical and each gene had a single intron. All the C. intestinalis vanabins were cysteine-rich, and the repetitive pattern of cysteines closely resembled that of A. sydneiensis samea vanabins. Using immobilized metal ion affinity chromatography (IMAC), we found that a recombinant protein of at least one of the C. intestinalis vanabins (CiVanabin5) bound to vanadium(IV) ions.

Chloride channel in vanadocytes of a vanadium-rich ascidian Ascidia sydneiensis samea.

Ascidians, so-called sea squirts, can accumulate high levels of vanadium in the vacuoles of signet ring cells, which are one type of ascidian blood cell and are also called vanadocytes. In addition to containing high concentrations of vanadium in the +3 oxidation state, the proton concentrations in vanadocyte vacuoles are extremely high. In order to elucidate the entire mechanism of the accumulation and reduction of vanadium by ascidian vanadocytes, it is necessary to clarify the participation of anions, which might be involved as counter ions in the active accumulation of both vanadium and protons. We examined the chloride channel, since chloride ions are necessary for the acidification of intracellular vesicles and coexist with H(+)-ATPase. We cloned a cDNA encoding a chloride channel from blood cells of a vanadium-rich ascidian, Ascidia sydneiensis samea. It encoded a 787-amino-acid protein, which showed striking similarity to mammalian ClC3/4/5-type chloride channels. Using a whole-mount in situ hybridization method that we developed for ascidian blood cells, the chloride channel was revealed to be transcribed in vanadocytes, suggesting its participation in the process of vanadium accumulation.

Expressed sequence tag analysis of blood cells in the vanadium-rich ascidian, Ascidia sydneiensis samea--a survey of genes for metal accumulation.

Some species in the family Ascidiidae accumulate vanadium at concentrations in excess of 350 mM, which corresponds to about 10(7) times that found in seawater. The vanadium ions are stored in vacuoles located within vanadium-containing blood cells, vanadocytes. To investigate the phenomenon, an expressed sequence tag analysis (EST) of a cDNA library of Ascidia sydneiensis samea blood cells was carried out. Three hundred clones were obtained and sequenced by EST analysis. A similarity search revealed that 158 of the clones (52.7%) were known genes, and 142 of the clones (47.3%) did not have any similarity to genes registered in the SwissProt database. According to the functions of their genes the identified EST clones were categorized into eight types of clones; these consisted of genes; metal-related proteins (29 clones), signal transduction (22 clones), protein synthesis (17 clones), nuclear proteins (17 clones), cytoskeleton and motility (14 clones), energy conversion (3 clones), hypothetical proteins (11 clones), and others (45 clones). The ferritin homologue has a high degree of similarity to that of mammals; the iron-binding sites of ferritin are well conserved including His-118 which is important for capturing Fe(2+), also works as a ligand for VO(2+).