Shoji Nakai

Role of b-lymphocytes in the immunopharmacological effects of a traditional chinese medicine, xiao-chai-hu-tang (shosaiko-to)

We previously reported that a traditional Chinese medicine, Xiao-chai-hu-tang (Japanese name: Shosaiko-to), induced interferon (IFN) activity in the serum of mice after intraperitoneal (i.p.) administration. In the present study in which murine spleen cells were cultured in vitro with Shosaiko-to, B-cells isolated by anti-immunoglobulin-coated plates were confirmed to generate IFN in response to Shosaiko-to stimulation. IFN activity was induced in the serum after i.p. administration of Glycyrrhizae radix, Scutellariae radix, Bupleuri radix and Pinelliae tuber which are included in Shosaiko-to as its constituent. Such an IFN-inducing activity was confirmed to exist in methanol-insoluble fractions of these extracts derived from Shosaiko-to and these constituents but not in methanol-soluble fractions. These four extracts as well as Shosaiko-to, induced interleukin 6 (IL-6) in the serum after the administration. In in vitro stimulation of spleen cells, Shosaiko-to and extracts of Glycyrrhizae radix, Bupleuri radix and Pinelliae tuber showed mitogenic activity, but an extract of Scutellariae radix with in vivo IFN-inducing activity did not. B-cells appear to participate in the immunopharmacological effects of Shosaiko-to through mitogenic activity, IFN induction and the effect of IL-6.

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.

Combined treatment of autoimmune MRL/MP-lpr/lpr mice with a herbal medicine, Ren-shen-yang-rong-tang (Japanese name: Ninjin-Youei-To) plus suboptimal dosage of prednisolone

Therapeutic effects of combined treatment with a Chinese medicine prescription, Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to, NYT) and suboptimal doses of prednisolone (PSL) on pathological findings of autoimmune-prone MRL/lpr mice were examined. Six-week-old MRL/lpr mice were treated orally with 1000 mg/kg of NYT, 0.5 or 2 mg/kg of PSL, 1000 mg/kg of NYT plus 0.5 or 2 mg/kg of PSL (combined treatment) or solvent only (control) six times per week. The rates of signs and symptoms of autoimmune disease (lymphadenopathy, proteinuria, dermatitis, loss of hair) were suppressed significantly in groups given PSL (2 mg/kg) alone, NYT alone and combined treatment with PSL (2 mg/kg) plus NYT (1000 mg/kg) compared with control, respectively, whereas treatment with PSL (0.5 mg/kg) alone did not inhibit their occurrence. ConA response and IL-2 production were also improved significantly in lymphocytes of mice given the combined treatment. Interestingly, treatment with NYT alone enhanced further the augmented IFN-gamma production in MRL/lpr mice but the combined treatment suppressed such an augmented production. The combined treatment dramatically reduced the level of anti-DNA antibodies in serum of MRL/lpr mice. By contrast, NYT alone treatment had no effect on autoantibodies production. These results suggest that combined treatment with NYT plus a suboptimal dose of PSL could be effective for systemic lupus erythematosus without severe side-effects.

Treatment effect of a traditional Chinese medicine, Ren-shen-yang-rong-tang (Japanese name: Ninjin-Youeito), on autoimmune MRL/MP-lpr/lpr mice

Abstract Autoimmune MRL /lpr mice were i.p. treated with 200 mg/kg Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to, NYT), a traditional Chinese herbal medicine (Japanese name: Kampo), from 8 weeks of age every 3 days before the onset of autoimmune disease. Compared to age-matched control MRL /lpr mice, the serum IL-6 concentration in NYT-treated mice was decreased, their serum IFN-γ concentration was increased, and the proliferative responses of whole and enriched CD4 + cells in their spleen and lymph nodes stimulated with ConA in vitro were restored. FACS analysis revealed that the rate of decreased CD4 + CD8 + T-cell population in the thymus was decreased in MRL /lpr mice but recovered by NYT treatment. Further, adult thymectomized (ATX) MRL /lpr mice were treated with 200 mg/kg NYT similarly. NYT treatment prolonged the survival of sham-operated MRL /lpr mice and ameliorated their proteinuria but did not improve such autoimmune manifestations in ATX-MRL /Ipr mice. These findings suggest that NYT plays an important role in the abrogation of autoimmune-prone T cell differentiation and that the therapeutic effect of NYT is dependent on the thymus in MRL /Ipr mice.

An extract of seeds fromAeginetia indica L., a parasitic plant, induces potent antigen-specific antitumor immunity in Meth A-bearing BALB/c mice

SummaryThe antitumor activity of an extract of seeds fromAeginetia indica L., a parasitic plant, was investigated. BALB/c mice, inoculated i.p. 1 × 105 syngeneic Meth A tumor cells, were administered 2.5 mg/kgA. indica extract i.p. every 2 days from day 0. The untreated mice died of an ascitic form of tumor growth within 21 days, whereas all the treated mice completely recovered from tumor challenge without any side-effects. The extract did not exert direct cytotoxic activity against Meth A in vitro. Mice that survived after the first challenge as a result ofA. indica treatment overcame the rechallenge with homologous Meth A without additional administration of the extract. On the other hand, those mice could not survive after rechallenge with Meth 1 tumor cells, which were also established in BALB/c mice but were different in antigenicity from Meth A, suggesting the development of antigen-specific concomitant immunity in theA. indica-cured mice. In the induction phase of antitumor resistance in this system, CD4+ T cells appeared to be the main contributors, since in vivo administration of anti-CD4 mAb completely abolished such resistance. In contrast, anti-CD8 mAb administration did not influence the effect ofA. indica. The importance of CD4+ T cells in antitumor immunity was again clarified by Winn assay; that is, spleen and lymph node cells depleted of CD4+ T cells in vitro prior to assay abolished antitumor activity on co-grafted Meth A tumor cells in vivo.

Combined treatment with Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to) plus prednisolone on adjuvant-induced arthritis in Lewis rat

The effects of combined treatment with 250 mg/kg Ren-shen-yang-rong-tang (Japanese name: Ninjinyouei-to, NYT) plus 4 mg/kg, an average dosage, or 0.2 mg/kg, a suboptimal dosage, of prednisolone (PSL) on adjuvant-induced arthritis in Lewis rats were investigated using two treatment schedules. The agents were administered orally every day from day 0 to 21 (schedule A), or from day -7 to 21 (schedule B) after adjuvant injection. PSL treatment (4 mg/kg) obviously inhibited paw swelling due to non-immune inflammation, diminished the weights of the thymus, spleen, adrenals and iliac lymph nodes, and suppressed the increment of serum interleukin (IL)-6 concentration in both schedules compared to controls. NYT treatment alone inhibited paw swelling due to immune inflammation, diminished the weight of the adrenals and decreased IL-6 concentration only in schedule A. Combined treatment with NYT plus PSL (4 mg/kg) showed: (1) a superior effect to that of PSL on paw swelling in the uninjected hind foot, especially in schedule B, (2) a tendency to diminish adrenal weight in schedule A and the weights of all four organs in schedule B compared with PSL treatment alone, and (3) a suppressive effect on IL-6 concentration weaker than that of PSL alone in schedule B. The suppressive effect of combined treatment with NYT plus PSL (0.2 mg/kg) on paw swelling was significantly stronger compared with either NYT or PSL treatment alone in schedule B. Although this dose of PSL had no influence upon the IL-6 concentration, the combined treatment or NYT alone increased the IL-6 concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Combined treatments with Ninjin-youei-to (Ren-shen-yang-rong-tang) plus a suboptimal dose of prednisolone on autoimmune nephritis in MRL\lpr mice

MRL/lpr mice suffer from a systemic lupus erythematosus-like autoimmune disease. We studied the effects of oral treatments with Ninjin-youei-to (NYT, Ren-shen-yang-rong-tang, 1000 mg/kg/day), a suboptimal dose (2 mg/kg/day) of prednisolone(PSL) and their combination on nephritis in MRL/lpr mice. Treatments with NYT or PSL alone inhibited the development of proteinuria and prolonged survival. The combined treatment reduced the incidence of proteinuria and prolonged survival. In histological analysis, NYT treatments decreased the degree of mesangial proliferative glomerulonephritis and infiltration of mononuclear cells in the kidneys. PSL treatment was effective in reducing periglomerular nephritis and vasculitis in addition to such effects as NYT and NYT plus PSL treatment was more effective than PSL alone. The active form of TGF-beta was reduced in NYT and PSL-treated mouse serum, and the combined treatments further suppressed it. However, the treatment with NYT alone did not induce a decrease in the latent form of TGF-beta. The effect of NYT can be assumed to be different from an immunosuppressive effect of PSL. Therefore, the combined treatment with NYT and PSL can be expected to be more useful for the therapy of autoimmune disease such as nephritis, compared with NYT or PSL alone treatments.

Comparison of Immunological Effects of Cholera Toxin on Autoimmune MRL/lpr and BXSB Mice

MRL/lpr and BXSB mice were treated weekly or biweekly with cholera toxin (CT) in intravenous dose of 2 μg/mouse. CT treatment notably alleviated proteinuria in MRL/lpr mice, but did not influence the course of lupus nephritis in BXSB male mice. Flow cytometric analysis showed that anomalous B220+ T cells in spleen and thymus were reduced in CT‐treated MRL/lpr mice while no significant change in lymphocyte populations was induced in BXSB male mice by this treatment. The suppressive effect of CT treatment on Con A response and the augmentative action on LPS response were observed in MRL/lpr mice. The latter may reflect increased B cells in relative number in the peripheral lymphoid organs. Mitogenic responses in CT‐treated BXSB male mice remained unchanged in comparison with those of untreated group. Increased production of IL‐6 by spleen cells was demonstrated in MRL/lpr mice treated with CT while in BXSB mice the level of IL‐6 was not changed by the treatment with CT. Production of IFNγ was suppressed by CT treatment in both strains of mice. This may be attributed to the inhibitory effect of CT on IFNγ‐producing Th1 cells as reported previously (Munoz et al, J. Exp. Med. 172: 95–103, 1990). However, CT treatment did not inhibit anti‐DNA antibody production in BXSB mice, whereas the autoantibodies were markedly decreased in MRL/lpr mice treated with CT.