Induction of interferon after administration of a traditional Chinese medicine, xiao-chai-hu-tang (shosaiko-to).

We examined the ability of a traditional chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to) to induce IFN in mice. A maximum activity (105 units/ml) of interferon (IFN) appeared in the serum of mice 16 h after intraperitoneal (i.p.) treatment with 250 mg/kg of shosaiko-to. Addition of polymyxin B did not abrogate the ability of shosaiko-to to induced serum IFN. The IFN was identified as IFN-alpha/beta by neutralizing test using anti-IFN alpha/beta antibodies. Pretreatment of mice with carrageenan suppressed the IFN induction by shosaiko-to, whereas the IFN induction by shosaiko-to was impaired neither in mice treated with anti-asialo-GM1 antibody nor in T-cell-deficient athymic nude mice. IFN was produced in vitro by spleen cells obtained from shosaiko-to treated mice. Moreover, spleen cells from untreated mice could also produce IFN when they were cultured with shosaiko-to. Additionally, serum IFN was also induced by the adoptive transfer of spleen cells from shosaiko-to treated mice to normal mice. On the other hand, peroral administration of shosaiko-to also induced IFN-alpha/beta in the serum. While IFN activity induced by i.p. administration of shosaiko-to declined after repeated treatments, the activity induced by its peroral administration did not decline during a long term treatment. These results showed that shosaiko-to is an IFN-alpha/beta inducer capable of repeated peroral administration.     

Contribution of Cytokines to Time-Dependent Augmentation of Resistance Against Listeria Monocytogenes After Administration of a Traditional Chinese Medicine,Xiao-Chai-Hu-Tang (Japanese Name: Shosaiko-To)

Abstract

The augmentation of resistance against Listeria monocytogenes after an intraperitoneal (ip) administration of shosaiko-to in mice was shown to depend on the time interval between the treatment and the infection. A maximal effect was expressed in mice treated 4 days before ip infection. The time dependent resistance correlated to the accumulation of macrophages in the peritoneal cavity just before the infection, but not to bactericidal activity as judged by the fact that peritoneal macrophages from untreated mice and those from mice treated with shosaiko-to 4 days before showed a high bactericidal activity of the same degree. Resistance to the infection in untreated mice may be attributable to newly accumulating macrophages with a low level of bactericidal activity, but not to resident macrophages with a high level of the activity. After intravenous infection, on the other hand, a maximal effect was expressed in mice treated with shosaiko-to 2 days before. The resistance correlated to accumulation of macrophages and bactericidal activity in the spleen just before the infection. Participation of cytokines in an augmenting effect of shosaiko-to on protection against the infection was examined. Shosaiko-to induced a transient elevation of serum CSF activity that was maximal at 3 hours after the administration in uninfected mice, though it did not augment the CSF activity induced by the infection. The elevation of CSF activity may induce accumulation of macrophages with a high level of bactericidal activity in the spleen 2 days after administration of shosaiko-to and then in the peritoneal cavity 4 days after administration. IFN-γ and TNF-α did not participate in the effect because administration of anti-IFN-γ or anti-TNF-α just before administration of shosaiko-to or just before infection did not abrogate the inhibitory effect of shosaiko-to on the bacterial growth in the early stage of infection. Shosaiko-to also induced an increase of CFUm number in the spleen. The effect may contribute to the augmentation of resistance in the late stage of infection by differentiating to mature macrophages.     

Activation of murine peritoneal macrophages by intraperitoneal administration of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to)

Macrophage activation by a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), was investigated. Intraperitoneal (i.p.) administration of shosaiko-to into (BALB/c x DBA/2)F1 mice resulted in marked activation of macrophages with respect to phagocytic and lysosomal enzyme activities (acid phosphatase and N-acetyl-beta-D-glucosaminidase) compared with the control. The maximal responses were induced by an i.p. injection of 3 mg shosaiko-to 4 days previously. Enhanced activities induced by shosaiko-to were also seen in C3H/HeJ mice, which is a non-responder strain to bacterial lipopolysaccharide (LPS). Significant macrophage accumulation in the peritoneal cavity and increased lysosomal enzyme activities were observed in mice injected with shosaiko-to. Shosaiko-to exhibited significant cytostasis-inducing activity. In addition, the administration of shosaiko-to led to a moderate expression of Ia antigen on the surface of peritoneal macrophages. These results suggest that shosaiko-to is a potent macrophage activator.     

Contribution of cytokines to time-dependent augmentation of resistance against Listeria monocytogenes after administration of a traditional Chinese medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to)

The augmentation of resistance against Listeria monocytogenes after an intraperitoneal (ip) administration of shosaiko-to in mice was shown to depend on the time interval between the treatment and the infection. A maximal effect was expressed in mice treated 4 days before ip infection. The time dependent resistance correlated to the accumulation of macrophages in the peritoneal cavity just before the infection, but not to bactericidal activity as judged by the fact that peritoneal macrophages from untreated mice and those from mice treated with shosaiko-to 4 days before showed a high bactericidal activity of the same degree. Resistance to the infection in untreated mice may be attributable to newly accumulating macrophages with a low level of bactericidal activity, but not to resident macrophages with a high level of the activity. After intravenous infection, on the other hand, a maximal effect was expressed in mice treated with shosaiko-to 2 days before. The resistance correlated to accumulation of macrophages and bactericidal activity in the spleen just before the infection. Participation of cytokines in an augmenting effect of shosaiko-to on protection against the infection was examined. Shosaiko-to induced a transient elevation of serum CSF activity that was maximal at 3 hours after the administration in uninfected mice, though it did not augment the CSF activity induced by the infection. The elevation of CSF activity may induce accumulation of macrophages with a high level of bactericidal activity in the spleen 2 days after administration of shosaiko-to and then in the peritoneal cavity 4 days after administration. IFN-gamma and TNF-alpha did not participate in the effect because administration of anti-IFN-gamma or anti-TNF-alpha just before administration of shosaiko-to or just before infection did not abrogate the inhibitory effect of shosaiko-to on the bacterial growth in the early stage of infection. Shosaiko-to also induced an increase of CFUm number in the spleen. The effect may contribute to the augmentation of resistance in the late stage of infection by differentiating to mature macrophages.     

Protective Effect of a Traditional Chinese Medicine, Xiao-Chai-Hu-Tang (Japanese Name:Shosaiko-To), On Pseudomonas Aeruginosa Infection in Mice

Survival of mice after intraperitoneal (ip) or intraveneous (iv) infection with Pseudomonas aeruginosa was augmented in the mice that had been pretreated ip with a Chinese traditional herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to) 6 hours or 4 days previously. 1) The pretreatment with shosaiko-to 6 hours previously induced an accumulation of polymorphonuclear leukocytes (PMN) in the peritoneal cavity, and its protective effect against ip infection was not impaired by treatment with carrageenan, a macrophage blocking agent. These results suggested that the protective effects of shosaiko-to against P. aeruginosa infection depended mainly on PMN in mice preteated at this timing. 2) The pretreatment with shosaiko-to 4 days previously induced an accumulation of macrophages showing an augmented phagocytosis of P. aeruginosa in vitro in the presence of immune serum, and its protective effect against P. aeruginosa was impaired by treatment with carrageenan. In addition, the pretreatment with shosaiko-to accelerated the bacterial clearance from the blood. The sera obtained from mice treated with shosaiko-to 4 days previously showed a high titer of antibody specific to P. aeruginosa. When this sera was transferred to naive mice, these recipients showed an accelerated bacterial clearance and an increased survival to challenge infection with P.aeruginosa. These results suggested that protective effects of shosaiko-to against P. aeruginosa infection at this timing depended on cooperation of macrophages and antibody which produced by stimulation of shosaiko-to, a polyclonal B cell activator. Such polyclonal antibodies were also effective on protection against encapsulated Klebsiella pneumoneae to which antibody was essential in the expression of resistance. These results suggested that shosaiko-to could augment nonspecific resistance to a variety of bacteria to which antibody plays an effective role.     

Accumulation of Immature B and Null Lymphocytes in the Periphery After Intraperitoneal Administration of Traditional Chinese Medicine, Xiao-Chai-Hu-Tang (Japanese Name : Shosaiko-to)

Accumulation of lymphocytes after an intraperitoneal (ip) injection of a traditional Chinese herb medicine, XIAO-CHAI-HU-TANG (Japanese name : shosaiko-to), was investigated. Shosaiko-to induced marked accumulation of lymphocytes rather than macrophages in the peritoneal cavity of ICR mice, whereas various kinds of irritants, e.g. proteose-pepton, Escherichia coli lipopoly-saccharide (LPS), OK-432 and Corynebacterium parvum, induced preferential accumulation of macrophages rather than lymphocytes. By means of analysis using two-color fluorescence-activated cell sorter (FACS), it was revealed that the increased lymphocyte subpopulations not only in the peritoneal cavity but also in the spleen of C3H/He mice by the injettion of shosaiko-to were comprised of both immature B(IgM⁺ and IgD^-) and null (thyl^- and Ig^-) cells. This effect of shosaiko-to was observed in other C5 normal strains, C3H/HeJ (LPS-nonresponder), C57BL/6, BALB/c and athymic nu/nu (ICR background) mice, but not in C5 deficient strains, AKR/J, A/J and DBA/2 mice, indicating that the accumulation of immature B and null cells in the periphery induced by shosaiko-to is closely related to the complement system.     

Role of macrophages in serum colony-stimulating factor induction by Lactobacillus casei in mice.

Heat-killed Lactobacillus casei YIT9018 (LC9018), when injected intravenously into mice at a dose of 4 to 40 mg/kg, induced the production of serum colony-stimulating factor (CSF). Since this induction was observed in both C3H/HeJ and C3H/HeN mice, LC9018 was considered to act differently from lipopolysaccharide. The amount of serum CSF induced by LC9018 in nude mice and whole-body-X-ray-irradiated mice was similar to that in control mice, but the induction of serum CSF was suppressed by the previous administration of carrageenan, indicating that macrophages, but not T cells, were responsible for serum CSF induction by LC9018. To determine whether macrophages themselves produce CSF or help other cells produce CSF in response to LC9018, we prepared adherent cells from the peritoneal cavity of normal mice and examined CSF activity in their conditioned media. Peritoneal adherent cells did not produce CSF without LC9018, but when cultivated with 1 mg of LC9018 per ml, they produced CSF at the same time that serum CSF was induced after the intravenous administration of LC9018. Additionally, in vitro-induced CSF formed macrophage, granulocyte, and mixed colonies, as serum CSF did. CSF production by peritoneal adherent cells was completely inhibited by cycloheximide (50 micrograms/ml), and neither the elimination of T cells from the peritoneal adherent cells by treating them with anti-Thy-1.2 antibody plus complement nor the addition of T cells affected CSF production. These results suggest that heat-killed LC9018 induces serum CSF in mice via direct stimulation of macrophages to produce CSF de novo.     

Functional Maturation of Immature b Cells Accumulated in the Periphery by an Intraperitoneal Administration of a Traditional Chinese Medicine, Xiao-Chai-Hu-Tang (Japanese Name: Shosaiko-To)

We found that an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), induced accumulation of B lymphocytes (sIgM⁺) in the peritoneal cavity and spleen. 1) Cell surface marker analysis by a fluorescence-activated cell sorter (FACS) demonstrated that the accumulated B cells on day 4 or 7 after shosaiko-to administration (early phase) were composed mainly of sIgM⁺ IgD^- cells and suggested that these B cells maturated into sIgM⁺ IgD⁺ cells on days 10 or 14 (late phase). Relative decrease of IgM⁺ IgD⁺cells at early phase was more profound in peritoneal cells (PC) than in spleen cells. 2) With respect to spleen lymphocytes, antibody responses to a thymus-independent (TI) antigen of type 2 (trinitrophenylated Ficoll) and a thymus-dependent (TD) antigen (sheep erythrocyte) were enhanced at late phase but not at early phase. In contrast, responses to trinitrophenylated lipopolysaccharide (TNP-LPS) as a TI-1 antigen and LPS as a B cell mitogen or a polyclonal B cell activator were enhanced markedly at early phase but declined at late phase. 3) With respect to peritoneal lymphocytes, responses to LPS were suppressed at early phase but recovered at late phase. Enhanced responses to TI and TD antigen at late phase in spleen lymphocytes and suppressed response to LPS at early phase in peritoneal lymphocytes may be explained by increases of IgM⁺ IgD⁺ mature B cells and IgM⁺ IgD^- immature B cells, respectively, at those times. Enhanced responses to TI-1 or LPS in spleen lymphocytes at early phase may be explained by elevated sensitivity of IgM⁺ IgD⁺ cells which reside in the spleen before shosaiko-to administration and receive the direct stimulation by shosaiko-to, or by acquired responsiveness of IgM⁺ IgD^- cells which migrate after stimulation with shosaiko-to.     

Functional maturation of immature B cells accumulated in the periphery by an intraperitoneal administration of a traditional Chinese medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to)

We found that an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), induced accumulation of B lymphocytes (sIgM+) in the peritoneal cavity and spleen. 1) Cell surface marker analysis by a fluorescence-activated cell sorter (FACS) demonstrated that the accumulated B cells on day 4 or 7 after shosaiko-to administration (early phase) were composed mainly of sIgM+IgD- cells and suggested that these B cells maturated into sIgM+IgD+ cells on days 10 or 14 (late phase). Relative decrease of IgM+IgD+ cells at early phase was more profound in peritoneal cells (PC) than in spleen cells. 2) With respect to spleen lymphocytes, antibody responses to a thymus-independent (TI) antigen of type 2 (trinitrophenylated Ficoll) and a thymus-dependent (TD) antigen (sheep erythrocyte) were enhanced at late phase but not at early phase. In contrast, responses to trinitrophenylated lipopolysaccharide (TNP-LPS) as a TI-1 antigen and LPS as a B cell mitogen or a polyclonal B cell activator were enhanced markedly at early phase but declined at late phase. 3) With respect to peritoneal lymphocytes, responses to LPS were suppressed at early phase but recovered at late phase. Enhanced responses to TI and TD antigen at late phase in spleen lymphocytes and suppressed response to LPS at early phase in peritoneal lymphocytes may be explained by increases of IgM+IgD+ mature B cells and IgM+IgD- immature B cells, respectively, at those times. Enhanced responses to TI-1 or LPS in spleen lymphocytes at early phase may be explained by elevated sensitivity of IgM+IgD+ cells which reside in the spleen before shosaiko-to administration and receive the direct stimulation by shosaiko-to, or by acquired responsiveness of IgM+IgD- cells which migrate after stimulation with shosaiko-to.     

Protective Effect of a Traditional Chinese Medicine,Xiao-Chai-Hu-Tang (Japanese Name:Shosaiko-To), On Pseudomonas Aeruginosa Infection in Mice

Abstract

Survival of mice after intraperitoneal (ip) or intraveneous (iv) infection with Pseudomonas aeruginosa was augmented in the mice that had been pretreated ip with a Chinese traditional herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to) 6 hours or 4 days previously. 1) The pretreatment with shosaiko-to 6 hours previously induced an accumulation of polymorphonuclear leukocytes (PMN) in the peritoneal cavity, and its protective effect against ip infection was not impaired by treatment with carrageenan, a macrophage blocking agent. These results suggested that the protective effects of shosaiko-to against P. aeruginosa infection depended mainly on PMN in mice preteated at this timing. 2) The pretreatment with shosaiko-to 4 days previously induced an accumulation of macrophages showing an augmented phagocytosis of P. aeruginosa in vitro in the presence of immune serum, and its protective effect against P. aeruginosa was impaired by treatment with carrageenan. In addition, the pretreatment with shosaiko-to accelerated the bacterial clearance from the blood. The sera obtained from mice treated with shosaiko-to 4 days previously showed a high titer of antibody specific to P. aeruginosa. When this sera was transferred to naive mice, these recipients showed an accelerated bacterial clearance and an increased survival to challenge infection with P.aeruginosa. These results suggested that protective effects of shosaiko-to against P. aeruginosa infection at this timing depended on cooperation of macrophages and antibody which produced by stimulation of shosaiko-to, a polyclonal B cell activator. Such polyclonal antibodies were also effective on protection against encapsulated Klebsiella pneumoneae to which antibody was essential in the expression of resistance. These results suggested that shosaiko-to could augment nonspecific resistance to a variety of bacteria to which antibody plays an effective role.     

Protective Effect of a Traditional Chinese Medicine, Xiao-Chai-Hu-Tang (Japanese Name: Shosaiko-To), On Listeria Monocytogenes Infection in Mice

Lethal effect of Listeria monocytogenes (L. monocytogenes) in mice was prevented by an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), 4 days before ip bacterial infection. The numbers of bacteria in the peritoneal cavity and liver were smaller in shosaiko-to-treated mice from one day after the infection. Macrophage accumulation in the peritoneal cavity after ip inoculation of L. monocytogenes was observed in both untreated and shosaiko-to-treated mice. Although rates of such increases were almost the same between both groups, the absolute number of macrophages was larger in shosaiko-to-treated than in untreated mice because of a higher level of the macrophage number at 4 days after ip injection of shosaiko-to. In untreated mice, bactericidal activity of peritoneal macrophages decreased from one day to 3 days after ip injection of killed L. monocytogenes. Such an activity was maintained at the same level from 1 to 3 days in shosaiko-to-treated mice. Augmented accumulation of macrophages and maintenance of their bactericidal activity may be main mechanisms of the augmented resistance in shosaiko-to-treated mice. Augmented resistance against bacterial growth in the thigh muscle in ip shosaiko-to-treated mice may be caused by such mechanisms. The effect of shosaiko-to observed at an early stage of infection may be T cell-independent, since such an effect was observed in athymic nude mice and delayed footpad reaction could not be detected at such a timing in euthymic normal mice.     

Functional Maturation of Immature b Cells Accumulated in the Periphery by an Intraperitoneal Administration of a Traditional Chinese Medicine,Xiao-Chai-Hu-Tang (Japanese Name: Shosaiko-To)

Abstract

We found that an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), induced accumulation of B lymphocytes (sIgM⁺) in the peritoneal cavity and spleen. 1) Cell surface marker analysis by a fluorescence-activated cell sorter (FACS) demonstrated that the accumulated B cells on day 4 or 7 after shosaiko-to administration (early phase) were composed mainly of sIgM⁺ IgD^- cells and suggested that these B cells maturated into sIgM⁺ IgD⁺ cells on days 10 or 14 (late phase). Relative decrease of IgM⁺ IgD⁺cells at early phase was more profound in peritoneal cells (PC) than in spleen cells. 2) With respect to spleen lymphocytes, antibody responses to a thymus-independent (TI) antigen of type 2 (trinitrophenylated Ficoll) and a thymus-dependent (TD) antigen (sheep erythrocyte) were enhanced at late phase but not at early phase. In contrast, responses to trinitrophenylated lipopolysaccharide (TNP-LPS) as a TI-1 antigen and LPS as a B cell mitogen or a polyclonal B cell activator were enhanced markedly at early phase but declined at late phase. 3) With respect to peritoneal lymphocytes, responses to LPS were suppressed at early phase but recovered at late phase. Enhanced responses to TI and TD antigen at late phase in spleen lymphocytes and suppressed response to LPS at early phase in peritoneal lymphocytes may be explained by increases of IgM⁺ IgD⁺ mature B cells and IgM⁺ IgD^- immature B cells, respectively, at those times. Enhanced responses to TI-1 or LPS in spleen lymphocytes at early phase may be explained by elevated sensitivity of IgM⁺ IgD⁺ cells which reside in the spleen before shosaiko-to administration and receive the direct stimulation by shosaiko-to, or by acquired responsiveness of IgM⁺ IgD^- cells which migrate after stimulation with shosaiko-to.     

Protective Effect of a Traditional Chinese Medicine,Xiao-Chai-Hu-Tang (Japanese Name: Shosaiko-To), On Listeria Monocytogenes Infection in Mice

Abstract

Lethal effect of Listeria monocytogenes (L. monocytogenes) in mice was prevented by an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), 4 days before ip bacterial infection. The numbers of bacteria in the peritoneal cavity and liver were smaller in shosaiko-to-treated mice from one day after the infection. Macrophage accumulation in the peritoneal cavity after ip inoculation of L. monocytogenes was observed in both untreated and shosaiko-to-treated mice. Although rates of such increases were almost the same between both groups, the absolute number of macrophages was larger in shosaiko-to-treated than in untreated mice because of a higher level of the macrophage number at 4 days after ip injection of shosaiko-to. In untreated mice, bactericidal activity of peritoneal macrophages decreased from one day to 3 days after ip injection of killed L. monocytogenes. Such an activity was maintained at the same level from 1 to 3 days in shosaiko-to-treated mice. Augmented accumulation of macrophages and maintenance of their bactericidal activity may be main mechanisms of the augmented resistance in shosaiko-to-treated mice. Augmented resistance against bacterial growth in the thigh muscle in ip shosaiko-to-treated mice may be caused by such mechanisms. The effect of shosaiko-to observed at an early stage of infection may be T cell-independent, since such an effect was observed in athymic nude mice and delayed footpad reaction could not be detected at such a timing in euthymic normal mice.     

The water-soluble extract from cultured medium of Ganoderma lucidum (Reishi) mycelia (Designated as MAK) ameliorates murine colitis induced by trinitrobenzene sulphonic acid

Ganoderma lucidum Karst is well known as 'Reishi', a traditional food in China and Japan. It contains a polysaccharide component known to induce granulocyte macrophage colony-stimulating factor (GM-CSF) production from murine splenocytes. Moreover, GM-CSF may be a therapeutic agent for Crohn's disease. In this study, we investigated the water-soluble, polysaccharide components of Reishi (designated as MAK) in murine colitis induced by trinitrobenzene sulphonic acid (TNBS). We examined the concentration of GM-CSF in peritoneal macrophage cells (PMs) of C57BL/6 mice during in vitro and in vivo stimulation with MAK. After feeding with chow or MAK for 2 weeks, 2 mg of TNBS/50% ethanol was administered to each mouse. After 3 days of TNBS treatment, intestinal inflammation was evaluated, and mononuclear cells of the mesenteric lymph nodes (MLNs) and colon were cultured for ELISA. To determine the preventive role of GM-CSF, the mice were pre-treated with or without anti-GM-CSF antibody before TNBS administration. In vitro and in vivo MAK-stimulated PMs produced GM-CSF in a dose-dependent manner. Intestinal inflammation by TNBS was improved by feeding with MAK. MLNs of mice treated with TNBS produced IFN-γ, which was inhibited by feeding with MAK. In contrast, MLNs of mice treated with TNBS inhibited GM-CSF production, which was induced by feeding with MAK. The colon organ culture assay also revealed that IFN-γ was decreased and GM-CSF was increased by MAK. The preventive effect was blocked by the neutralization of GM-CSF. We concluded that the induction of GM-CSF by MAK may provide the anti-inflammatory effect.     

A traditional Chinese herbal medicine, ren-shen-yang-rong-tang (Japanese name: ninjin-yoei-to) augments the production of granulocyte-macrophage colony-stimulating factor from human peripheral blood mononuclear cells in vitro.

Ren-shen-yang-rong-tang (Japanese name: Ninjin-yoei-to, NYT) is a traditional Chinese herbal medicine. Leukocytosis and elevated levels of colony-stimulating factor (CSF) in peripheral blood were found previously after the administration of this compound to mice. In this study, human peripheral blood mononuclear cells (PBMC) were cultured in the presence of NYT in vitro, and the levels of granulocyte-macrophage CSF (GM-CSF) and granulocyte CSF (G-CSF) in the supernatant of cultured PBMC were measured using a sensitive enzyme-linked immunosorbent assays. NYT significantly (P less than 0.01) augmented GM-CSF production but not G-CSF production by PBMC in vitro.     

Effect of a traditional Chinese herbal medicine ren-shen-yang-rong-tang (Japanese name: ninjin-youei-to) on hematopoietic stem cells in mice

Cell dynamics after intraperitoneal (i.p.) and oral administration of a traditional herbal medicine, ren-shen-yang-rong-tang (Japanese name: ninjin-youei-to, NYT), were investigated. When NYT was injected i.p. into C3H/He mice, numbers of spleen and peritoneal cells significantly increased in a dose-dependent manner and showed high levels from 4 to 21 days. Two peaks in the total cell number were observed on days 7 and 14 in the peritoneal cavities and spleen of C3H/He mice administered NYT. A marked accumulation of PMN cells in the peritoneal cavity and spleen was detected at 7 days after injection. The numbers of macrophages and lymphocytes also increased by i.p. administration of NYT. The thymus cell number decreased transiently between 4 and 7 days and thereafter returned to the control level. No significant change in the cell number of lymph nodes was observed. Such cellular accumulation was also detected in C3H/HeJ mice, a nonresponder strain to bacterial endotoxin, and athymic nude mice. The activity of colony-forming units in the spleen (CFU-S) of C3H/He as well as C3H/HeJ mice was markedly augmented by i.p. administration of NYT. NYT induced significant CSF production as detectable by its activity in the sera. In addition, oral administration of NYT for 10 days induced a significant increment of peripheral leukocytes and spleen cells and enhanced CFU-S activity of bone marrow cells as induced by i.p. administration, indicating that NYT acts on hematopoietic stem cells capable of differentiating to lymphocytes, macrophages and PMN cells into the periphery.     

Augmentation of Host Resistance to Listeria Monocytogenes Infection by A Traditional Chinese Medicine, Ren-Shen-Yang-Rong-Tang (Japanese Name: Ninjin-Youei-To)

Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to, NIN), a traditional Chinese medicine, is a drug made of spray-dried powder of hot water extract obtained from twelve species of medical plants. An intraperitoneal (ip) injection with NIN 2 days before intravenous (iv) infection with Listeria monocytogenes (L. monocytogenes) accelerated elimination of viable bacteria in the spleen in the early stage of infection (from day 1) and protected mice from the lethal infection. It was suggested that the protective effect of NIN was mediated by the activation of nonimmune macrophages playing a principle role in resistance in the early stage of infection. Two days after ip injection with NIN just before infection, significantly increment in the number of monocytes in the peripheral blood was observed, though macrophage number in the spleen and their intracellular killing activity were unchanged. At 12 hours after infection with L. monocytogenes a significantly enhanced increase of splenic macrophage number was observed in NIN-treated mice, compared to controls. After ip injection of NIN, interleukin-1 (IL-1), IL-6 and granulocyte macrophage-colony stimulating factor (GM-CSF) became detectable in the serum or peritoneal cavity. These results suggested that NIN stimulated macrophage-precursor cells in the bone marrow via the production of IL-1, IL-6, GM-CSF by macrophages, accelerated the supply of peripheral macrophages, and such macrophages accumulated into the site of infection in the very early stage of infection. Similar protective effects of NIN were observed by oral administration for 7 days till 1 day before iv infection with L. monocytogenes.     

Compartmentalized transgene expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in mouse lung enhances allergic airways inflammation

To investigate the role of GM-CSF in asthmatic airways inflammation, we have targeted GM-CSF transgene to the airway cells in a mouse model of ovalbumin (OVA)-induced allergic airways inflammation, a model in which there is marked induction of endogenous IL-5 and IL-4 but not GM-CSF. Following intranasal delivery of a replication-deficient adenoviral gene transfer vector (Ad), transgene expression was found localized primarily to the respiratory epithelial cells. Intranasal delivery of 0.03 × 10⁹ plaque-forming units (PFU) of AdGM-CSF into naive BALB/c mice resulted in prolonged and compartmentalized release of GM-CSF transgene protein with a peak concentration of ≈ 80 pg/ml detected in bronchoalveolar lavage fluid (BALF) at day 7, but little in serum. These levels of local GM-CSF expression per se resulted in no eosinophilia and only a minimum of tissue inflammatory responses in the lung of naive mice, similar to those induced by the control vector. However, such GM-CSF expression in the airways of OVA-sensitized mice resulted in a much greater and sustained accumulation of various inflammatory cell types, most noticeably eosinophils, both in BALF and airway tissues for 15–21 days post-OVA aerosol challenge, at which times airways inflammation had largely resolved in control mice. While the levels of IL-5 and IL-4 in BALF and the rate of eosinophil apoptosis were found similar between different treatments, there was an increased number of proliferative leucocytes in the lung receiving GM-CSF gene transfer. Our results thus provide direct experimental evidence that GM-CSF can significantly contribute to the development of allergic airways inflammation through potentiating and prolonging inflammatory infiltration induced by cytokines such as IL-5 and IL-4.     

Endotoxins prevent murine IgE production,T(H)2 immune responses,and development of airway eosinophilia but not airway hyperreactivity

BACKGROUND: Contact with immunomodulatory factors, such as LPS, in early infancy is associated with decreased allergen sensitization. OBJECTIVE: We sought to study the effects of systemic or airway exposure with LPS on the development of allergen sensitization, eosinophilic airway inflammation, and increased in vivo airway reactivity (AR) in a mouse model. METHODS: BALB/c mice were systemically sensitized with ovalbumin (OVA) plus adjuvant on days 1 and 14 and challenged through the airways with allergen on days 34 to 36. We performed measurement of OVA-specific IgE serum levels, in vitro T(H)2 cytokine production, differential cell counts in bronchoalveolar lavage fluids, and assessment of in vivo AR to inhaled methacholine by means of barometric whole-body plethysmography. RESULTS: Systemic LPS administration before OVA sensitization reduced OVA-specific IgE serum levels (426 +/- 76 vs 880 +/- 104 U/mL, P <.01), T(H)2 cytokine production by splenic mononuclear cells (IL-4: 0.08 +/- 0.01 vs 0.17 +/- 0.01 ng/mL; IL-5: 1.98 +/- 0.52 vs 4.11 +/- 0.54 ng/mL; P <.01), and extent of airway eosinophilia (total cell counts: 93 vs 376 x 10(3)/mL; eosinophils: 23% vs 51%; P <.01) compared with that in OVA-sensitized mice. Local LPS administration to sensitized mice before airway allergen challenges particularly induced IFN-gamma production by peribronchial lymph node cells in vitro (1718 +/- 315 vs 483 +/- 103 ng/mL, P <.01) associated with reduced airway eosinophilia compared with that seen in OVA-sensitized mice. Development of increased AR was not affected by systemic or local LPS exposure. Inhibitory effects of LPS on allergen sensitization and eosinophilic airway inflammation were inhibited by administration of anti-IL-12 antibodies before LPS exposure. CONCLUSION: These data indicate that local and systemic application of LPS modulates systemic and local T(H)1/T(H)2 immune responses in a distinct but similarly IL-12-dependent mode.