Takako Kasahara

Role of endogenous interferon-γ on the enhancement of splenic NK cell activity by electroacupuncture stimulation in mice

Successive electro-acupuncture (EA) stimulation applied to bilateral anterior tibial muscles, where Zusanli (ST36) acupoints are located, once a day (30 min) for 3 successive days significantly enhanced splenic natural killer (NK) cell activity in BALB/c mice. The percentage of splenic NK cells, as measured by flow cytometry, was not affected in these mice. Interferon (IFN)-gamma level in splenic aqueous extract, prepared from the ST36 acupoint-stimulated mice, was significantly higher than that of the controls. In vivo treatment with neutralizing monoclonal antibody against mouse IFN-gamma completely abrogated the increase in splenic NK cell activity induced by ST36 acupoint stimulation. The same stimulation also significantly increased the concentration of splenic beta-endorphin, which coincided with the significant increase in splenic IFN-gamma production. Pre-administration of 10 mg/kg naloxone before initiation of EA stimulation every day reduced the enhancements of NK cell activity and IFN-gamma level. These observations strongly suggest that endogenous IFN-gamma mediates the up-regulation of NK cell activity by EA stimulation at the ST36 acupoints. Furthermore, endogenous beta-endorphin secreted by EA stimulation also plays an important role in the up-regulation of NK cell function, which may be realized through regulating IFN-gamma production.

Suppressive effects of Tripterygium wilfordii Hook f., a traditional Chinese medicine, on collagen arthritis in mice.

The effect of chloroform extract of Tripterygium wilfordii Hook f. (TWH extract), a traditional immunosuppressive Chinese herb, on type II collagen (C II)-induced arthritis (CIA) in DBA/1J mice was studied. In the first set of experiments, we examined the effect of TWH extract on cellular immune responses to C II. As compared with mice treated with saline, TWH extract administered orally at doses of more than 400 microg kg(-1) once a day for 14 days inhibited the ability of inguinal lymph node cells to produce T cell cytokines interleukin-2 and interferon-gamma when the cells were obtained from mice 21 days after immunization and cultured in vitro with C II. Treatment with TWH extract also inhibited production of macrophage cytokines interleukin-1beta and tumor necrosis factor-alpha in response to in vitro stimulation of lymph node cells with C II. In the second part of the experiment, we evaluated the influence of TWH extract on the incidence and development of arthritis in murine CIA. Mice were immunized twice at a 3-week interval with bovine C II, with TWH extract being given orally once a day for 14 days with four different regimens. A 14-day course of TWH extract treatment at a daily dose of 400 microg kg(-1), which began on the day of the first C II immunization, suppressed the development of arthritis, as well as antibody production and delayed-type hypersensitivity to C II. Treatment with TWH extract, which started on the same day as the booster immunization, also resulted in inhibition of development of arthritis and of immune responses to C II. On the other hand, therapeutic administration with TWH extract did not affect the clinical course of the disease and the immune response to C II.

Inhibition of murine chronic graft-versus-host disease by the chloroform extract of Tripterygium wilfordii Hook f.

The effects of chloroform extract of Tripterygium Wilfordii Hook f (TWH extract) on chronic graft-versus-host disease (GVHD) were examined in a murine experimental model. Chronic GVHD was induced by intravenous transfer of parental DBA/2 spleen cells into unirradiated (C57BL/6 x DBA/2)F1 recipient mice. The effects of TWH extract on GVHD were assessed by measuring both the degree of splenomegaly and the total serum IgE levels 3 weeks after the cell transfer. Subcutaneous administration of TWH extract once a day for 3 weeks suppressed chronic GVHD in a dose-dependent manner. Significant suppression of splenomegaly was first noted in mice treated with 7.5 micrograms/kg of the agent. The maximum inhibition was observed when mice were treated with more than 10.0 micrograms/kg (but not 5.0 micrograms/kg) caused complete suppression of serum IgE hyperproduction. The ability of donor T cells purified from recipient spleen cells to produce interleukin 4 in response to stimulation with anti-CD3 monoclonal antibody was significantly abrogated when recipient mice were treated with 10.0 micrograms/kg of the agent. These results strongly suggest that TWH extract will be an addition to the cohort of immunosuppressive therapies used in solid organ and bone marrow transplantation.

Suppressive effect of acupuncture on delayed type hypersensitivity to trinitrochlorobenzene and involvement of opiate receptors

We reported previously that electroacupuncture (Acu) at an acu-point equivalent to GV-4 in mice either enhanced or suppressed the delayed type hypersensitivity (DTH) to 2,4,6-trinitrochlorobenzene (TNCB, picryl chloride) depending on the time of treatment. We report here the suppression of the efferent phase of DTH to TNCB by Acu in mice. In 7- to 9-week-old male BALB/c, C57BL/6 and ddY mice, significant suppression of the DTH was observed when Acu had been applied once per day for three consecutive days before TNCB challenge. When Acu had been applied a single, significant suppression also occurred. Application to another point (at a middle area of the femoral muscle) failed to suppress the DTH to TNCB. This Acu-evoked DTH suppression was blocked dose-relatedly by pretreatment of systemic naloxone hydrochloride, indicating that opioid receptor-mediated mechanisms are involved in this immune response.

Involvement of central opioidergic and nonopioidergic neuroendocrine systems in the suppressive effect of acupuncture on delayed type hypersensitivity in mice

The effect of a single treatment of electroacupuncture (Acu) at early or late stages of the efferent phase on 2, 4, 6-trinitrochlorobenzene (TNCB)-induced delayed type hypersensitivity (DTH) was studied in intact and hypophysectomized (HPX) mice. Acu (2.5 Hz, 15 min) applied to the acu-point equivalent to GV4 at 0, 3, 18 or 21 h after TNCB challenge induced significant suppression (45-73%) of the maximal extent of ear swelling at 24 h after TNCB challenge. An immunosuppressive and antiinflammatory drug, prednisolone 10 mg/kg i.p., also suppressed the DTH to the same extent. Pretreatment with intracisternal injection of naloxone hydrochloride (2 micrograms) significantly blocked the Acu-evoked DTH suppression when Acu treatment was done at 0 or 3 h. On the contrary, naloxone did not block the effect of Acu treatment given at 21 h. In order to examine the potential involvement of the pituitary in the suppression of DTH by Acu, the DTH reaction was examined in HPX mice. Acu failed to produce suppressive response in the HPX mice unless given at 0 h. These findings indicate that Acu treatment at acu-point GV4 during the efferent phase of induced DTH can suppress the DTH through central opioidergic or nonopioidergic systems. The pituitary is apparently pivotal in this immunosuppression and it is suggested that the DTH suppression by Acu may be mediated via activation of the neuroendocrine system.

Antipyretic action of peripheral stimulation with electroacupuncture in rats

The present study was designed to investigate the antipyretic action of peripheral stimulation with electroacupuncture (EA) in SD rats. EA stimulation was applied for 30 min to the peripheral muscle where the equivalent Quchi (LI11) acupoint is located. We first examined the effects of EA stimulation on fever induced by either lipopolysaccharide (LPS), interleukin-1 beta (IL-1 beta) or prostaglandin E2 (PGE2). Intraperitoneal injection of LPS at a dose of 100 micrograms/kg caused a high rectal temperature, which was suppressed by EA stimulation. EA stimulation also inhibited the development of fever induced by IL-1 beta injection either intravenously or into the preoptic area (POA). The rats that received administration of PGE2 into POA developed rapid and high fevers, which were attenuated by EA stimulation. In the second part of the experiment, we investigated the levels of cytokines and PGE2 during the development of fever. The concentrations of IL-6 and PGE2 but not IL-1 beta, in brain and serum were increased by intraperitoneal injection of 100 micrograms/kg LPS. EA stimulation reduced PGE2 levels in both brain and serum in LPS-injected rats, but failed to change the levels of IL-6. Intravenous injection of IL-1 beta produced high brain and serum PGE2 levels, which were also significantly suppressed by EA stimulation. The results suggest that EA stimulation possesses an antipyretic effect through the inhibition of the action of PGE2 in rats.

Suppressive activity of the chloroform extract of Tripterygium wilfordii Hook f on effector T cell activation during Hymenolepis nana infection in mice.

The chloroform extract of Tripterygium wilfordii Hook f (TWH extract) administered into mice daily at doses of 80.0 to 200.0 micrograms/kg (but not 40.0 micrograms/kg) caused suppression of protective immunity to Hymenolepis nana when the extract was injected subcutaneously during the induction phase of protective immunity. Daily administration of 200.0 micrograms/kg TWH extract, during the course of larval development from challenge, also suppressed protective immunity. Inhibition of protective immunity was only observed in mice that received TWH extract for 6 days at a daily dose of 200.0 micrograms/kg and were challenged 24 h after the final injection. TWH extract did not inhibit formation of effector cells that mediate delayed type hypersensitivity (DTH) to H. nana egg antigen when the extract was administered subcutaneously at a dose of 200.0 micrograms/kg/day for 5 days before cell preparation. However, TWH extract did inhibit DTH effector cell activation when cells prepared from infected, PBS-injected mice were transferred into 200.0 micrograms/kg TWH extract-treated recipient mice. These results strongly indicate that TWH extract cannot inhibit the generation of effector cells but will suppress their function in vivo.

The effect of acupuncture on natural killer cell activity

Abstract Successive electro-acupuncture (EA) stimulation applied to bilateral anterior tibial muscles, where Zusanli (ST36) acupoints are located, once a day (30 min) for 3 successive days significantly enhanced splenic natural killer (NK) cell activity in BALB/c mice. The percentage of splenic NK cells, as measured by flow cytometry, was not affected in these mice. Interferon (IFN)-γ level in splenic aqueous extract, prepared from the ST36 acupoint-stimulated mice, was significantly higher than that of the controls. In vivo treatment with neutralizing monoclonal antibody against mouse IFN-γ completely abrogated the increase in splenic NK cell activity induced by ST36 acupoint stimulation. The same stimulation also significantly increased the concentration of splenic β-endorphin, which coincided with a significant increase in splenic IFN-γ production. Pre-administration of 10 mg/kg naloxone before initiation of EA stimulation every day reduced the enhancement of NK cell activity and IFN-γ level. These observations strongly suggest that endogenous IFN-γ mediates the up-regulation of NK cell activity by EA stimulation at the ST36 acupoints. Furthermore, endogenous β-endorphin secreted by EA stimulation also plays an important role in the up-regulation of NK cell function, which may be realized through regulating IFN-γ production.

Effect of Single Moxibustion on Phagocytic Activity in Mice

Effects of moxibustion stimulation on the phagocytic activity of the reticuloendothelial system in ddY and ICR mice has been studied by using the carbon clearance methods. It was found that moxibustion stimulation induced the enhancement of the phagocytic activity with increased phagocytic indexes (K and a indexes) and lysosomal enzyme activities in mice peritoneal exudate cells and peritoneal macrophages. In addition, the increase of carbon uptake in the Kupffer cells of the liver after carbon injection can be seen by light microscopy when compared with that of nontreated mice. Few big holes on the cell surface of the macrophages obtained from the moxibustion mice were observed in scanning electron microscopical studies. These results suggest that the moxibustion treatment caused the enhancement of the host defence mechanisms in mice.

Inhibitory Effect of Electroacupuncture on Murine Collagen Arthritis and its Possible Mechanisms

The influence of electroacupuncture (EA), a traditional Chinese medical treatment, on type II collagen-induced arthritis (CIA) was examined in DBA/IJ mice in vivo. Mice were immunized intradermally twice at a 3-week interval with bovine type II collagen (C II). EA stimulation, begun on day 21 simultaneously with the second immunization, was applied at the acupoint equivalent to GV4 three times a week for 3 weeks. The results showed that EA delayed the onset, attenuated the severity of arthritis, and reduced the anti-collagen antibody level. Furthermore, we investigated the impact of EA on the productions of endogenous interleukin-1 beta (IL-1 beta) and prostaglandin E2 (PGE2), and the levels of IL-1 beta mRNA in splenocytes and synovial tissues from C II immunized mice on day 45 and cyclooxygenase-2 (COX-2) mRNA in lipopolysaccharide (LPS)-stimulated macrophages of normal mice by using reverse transcriptase-polymerase chain reaction (RT-PCR). EA stimulation significantly inhibited the concentrations of splenic endogenous IL-1 beta and serum PGE2. The expression of IL-1 beta mRNA in spleen cells was obviously down-regulated and that in synovial tissues was modestly affected by EA. COX-2 mRNA was highly expressed in cultured peritoneal macrophages when stimulated with LPS. Previous treatment with EA also reduced LPS-stimulated induction of COX-2 mRNA. These data suggest that EA has an inhibitory effect on murine CIA, and the partial mechanism of its therapeutic result may be attributed to inhibiting the productions of IL-1 beta and PGE2 by suppressing the IL-beta and COX-2 gene activations.