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.     

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.     

Thymus-dependent effects of a traditional chinese medicine,ren-shen-yang-rong-tang (Japanese name; Ninjin-Youei-To), in autoimmune MRL/MP-Ipr/Ipr mice

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.     

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.     

Recipient-mediated effect of a traditional chinese herbal medicine,Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to), on hematopoietic recovery following lethal irradiation and syngeneic bone marrow transplantation

Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to, NYT), a traditional Chinese herbal medicine, was evaluated for recipient-mediated effect on hematopoietic recovery in a murine model of syngeneic bone marrow transplantation (BMT). BALB/c recipient mice were preconditioned with a lethal total body irradiation (TBI) at a dose of 6.5 Gy and transplanted with syngeneic bone marrow (BM) cells. NYT treatments, given intraperitoneally (i.p.) once per day for 3 consecutive days in a dose of 0.625 mg, were performed either before or after TBI and BMT to assess any recipient-mediated effect of this compound. NYT pretreatment was as effective as NYT posttreatment in enhancing the total number of colony-forming unit erythroid (CFU-E) and colony-forming unit granulocyte-macrophage (CFU-GM) per marrow and spleen after TBI and BMT. NYT pretreatment caused a significant increase in marrow and splenic CFU-E and CFU-GM numbers over a prolonged period following TBI and BMT, and affected late-stage erythropoiesis (CFU-E) more profoundly than early-stage erythropoiesis (burst-forming unit erythroid, BFU-E). NYT pretreatment significantly accelerate recovery of not only erythrocyte and leukocyte counts but also platelet counts after transplantation with a limited number (1 × 10⁵) of BM cells. The same treatment, however, was significantly less effective in hematopoietic recovery after transplantation with a minimal number (1 × 10⁴) of BM cells, indicating that NYT accelerates recovery of donor-derived rather than recipient-derived cells. The data are consistent with the idea that NYT has an enhancing effect on hematopoiesis via the recipient microenvironment, and suggest that NYT may have an important role in the acceleration of hematopoietic recovery of donor-derived cells following BMT.     

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

MRL/Mp-lpr/lpr(MRL/lpr) mice were treated with a traditional Chinese herbal medicine, Ren-shen-yang-rong-tang (Japanese name: Ninjin-youei-to, NYT) intraperitoneally (i.p.) every 3 days or per os (p.o.) 6 times/week from before the onset of autoimmune disease (6 weeks of age). Fifty percent survival time was found in placebo-controlled male and female mice of 28 and 22 weeks of age, respectively. NYT-treatment markedly prolonged the survival time of MRL/lpr mice. That is, 50% survival time was 43 weeks in the i.p.-treated male mice and 30 weeks of age in the p.o.-treated female mice. Further, NYT-treatment significantly reduced occurence of thymic atrophy and prevented the anomalous accumulation of B220⁺ T-cells in lymphnode and spleen, both of which are characteristic in MRL/lpr mice. Moreover, grades of proteinuria were significantly reduced in both the i.p.- and p.o.-treated groups compared with the control groups. Such clinical benefit and increased survival time were interestingly not associated with the decrease in the level of autoantibodies.     

干细胞在心脏再生医学中的作用及中医药研究策略

背景：中医对心脏再生有其独特认识,但中医药介入干细胞与心脏再生医学还处于初步探索阶段。 目的：分析和总结干细胞与心脏再生相关的研究进展,对中医药如何介入心脏再生研究进行初步探讨。 方法：以关键词“stem cell”与 “cardiac regenerative medicine”检索PubMed数据库、关键词“干细胞”与“中医”检索CNKI中国期刊全文数据库2000/2010的相关文献,语言种类不限,对符合标准的文献证据进行综合提炼分析。保留其中44篇进行归纳总结。 结果与结论：分析表明干细胞应用于心脏再生医学的研究已广泛拓展,包括胚胎干细胞、骨髓干细胞、成体心脏干细胞等均显示了再生心肌的能力及其应用前景。中医对心脏再生的认识蕴含在阴阳观、五行母子规律及精气学说中。中医药要介入心脏再生医学,就需要对中医理论在心脏再生医学中的认识进行系统整理,深入挖掘中医复方,重视中药有效组分靶向干细胞及加强对有效方药作用机制的研究。     

Inhibition of eosinophil infiltration into the mouse peritoneal cavity by a traditional Chinese medicine, Bu-zhong-yi-qi-tang (Japanese name: Hochu-ekki-to)

Our previous study showed that the serum level of antigen-specific IgE antibodies in primary response was decreased by a traditional Chinese medicine, Bu-zhong-yi-qi-tang (Japanese name; Hochu-ekki-to, HOT). In this study, we examined inhibition of secondary IgE response and of eosinophil infiltration by HOT. BALB/c mice were intraperitoneally immunized with aluminum hydroxide adsorbed with DNP-KLH (DNP-KLH + alum) on day -14 and on day 0. In mice treated with HOT daily from day -14, the serum level of antigen-specific IgE antibodies after the secondary immunization was significantly decreased compared to that in mice not treated with HOT. Eosinophils increased in number after 6 and 24 hr, and CD4+ T cells in the peritoneal cavity increased in number 24 hr after the secondary immunization. HOT suppressed accumulation of eosinophils and CD4+ T cells in the peritoneal cavity. Furthermore, HOT suppressed the numbers of IL-4- and IL-5-producing cells 24 hr after the secondary immunization, but did not inhibit the number of IFN-gamma-producing cells. HOT also suppressed IL-5 mRNA expression. Furthermore, HOT also inhibited antigen-induced late-phase reaction (LPR) in the skin. These results suggested that HOT exhibited anti-allergic effects mainly by inhibiting Th2 cell responses.     

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.     

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.     

骨髓间充质干细胞对致敏小鼠造血干/祖细胞移植植入的影响

目的: 前期的研究已经证实致敏小鼠造血干/祖细胞移植植入失败率高。本研究拟通过骨髓间充质干细胞(MSCs)进行干预,观察能否提高造血干、祖细胞移植的植入率。方法: 应用贴壁培养法体外培养正常小鼠骨髓MSCs,并分为6个实验组,包括实验组1:d11 MSCs干预的致敏组;实验组2: d0 MSCs干预的致敏组;实验组3:d11和d0 2次MSCs干预的致敏组;实验组4: 无MSCs干预的致敏小鼠对照组;实验组5:无MSCs干预的正常小鼠(非致敏小鼠)移植对照组;实验组6:无MSCs干预的正常小鼠不移植对照组。观察指标包括生存分析、移植效果分析(血象改变、骨髓细胞恢复及嵌合分析等)和移植物抗宿主病(GVHD)检测,最终评估MSCs干预对各实验组异基因造血干/祖细胞移植植入率的影响效果。结果: 与对照组(实验组4、5、6)比较,MSCs干预(实验组1、2、3)在2次异基因脾细胞注射法致敏的动物模型进行异基因造血干/祖细胞移植时,未能促进骨髓造血干/祖细胞移植的植入,也未能延长致敏动物移植后的生存时间。结论: 体内应用1×10⁶ MSCs干预,未能促进2次异基因1×10⁶ C57BL/6小鼠脾细胞输注法建立的重度致敏模型异基因造血干/祖细胞移植的植入。     

Effect of traditional Chinese medicine (Sairei-to) on monoclonal antibody-induced proteinuria in rats

The effects of traditional Chinese medicine (Sairei-to) on monoclonal antibody (mAb) inducing proteinuria were examined. Four hundred, 200 and 100mglkg body weight (BW) of Sairei-to and phosphate-buffered saline (PBS) as a control were injected intraperitoneally into four groups of female Wistar rats every day from 5 days before intravenous injection of mAb to the end of the experimental period. The amount of urinary protein excretion was significantly suppressed in roughly a dose-dependent manner. For example, 116.6 ± 89.7mg/day of proteinuria was observed in control groups compared to 4.2 ± 15.2 mglday in the 400 mglkg BW of Sairei-to treated group 2 days after mAb injection (P < 0.01).
Statistically significant (P <0.01) differences were again observed in a repeat experiment (122.1 ± 53.7 vs 10.2 ± 10.1 mglday on the 2nd day) without affecting the glomerular filtration rate.
No significant difference was recognized between the total amount of mAb bound to glomeruli 1 h after mAb injection in Sairei-to-treated and non-treated rats, indicating that Sairei-to pretreatment has no effects on the number or quality of antigenic molecules.
The effect of Sairei-to on a non-immunological model of proteinuria was also examined. No significant reduction of proteinuria by similar Sairei-to treatment was observed in aminonucleoside of puromycin nephropathy.
The authors conclude that mAb-induced proteinuria in rats is significantly suppressed by the traditional Chinese medicine, Sairei-to.     

骨髓间质干细胞输注对外周血干细胞移植造血恢复的影响

目的:观察骨髓间质干细胞 (MSCs)输注对外周血干细胞移植 (PBSCT)后造血恢复的影响。方法:去脾小鼠经粒细胞集落刺激因子动员后收获外周血单个核细胞 (PBMC)和扩增培养的骨髓间质干细胞 (MSCs),移植给经放 /化疗预处理的BALB/c小鼠,数量分别为10⁶PBMC(PBSCT组)、10⁴MSCs和10⁶ PBMC(实验1组)、10⁶ MSCs和10⁶ PBMC(实验 2组),观察受体鼠 4周的生存率、骨髓有核细胞 (BMNC)、粒细胞巨噬细胞集落形成单位 (CFU-GM)、成纤维细胞集落形成单位 (CFU-F)、外周血白细胞 (WBC)计数等指标。结果:实验 2组的生存率、BMNC、CFU-GM、CFU-F显著高于PBSCT组,WBC计数恢复较PBSCT组快 (P <0.05);实验1组和PBSCT组比较,WBC计算恢复快,CFU-F产率高 (P <0.05)。结论:骨髓间质干细胞输注有促进外周血干细胞移植造血恢复的作用。     

Age-dependent abnormalities of hematopoietic stem cells in (NZW x BXSB)F1 mice

The (NZW x BXSB)F1 (W/BF1) mouse is known as an autoimmune-prone strain which develops lupus nephritis, thrombocytopenia due to platelet-specific autoantibodies, leukocytosis, and myocardial infarction. In this experiment, we investigated the age-dependent abnormalities of the hematopoietic stem cells (HSCs) and hematopoiesis in this mouse. White blood cell counts (especially Mac-1- or Gr-1-positive cells) in the peripheral blood of 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or normal mice. To investigate whether the abnormal hematopoiesis can be attributed to the HSCs of W/BF1 mice, colony-forming unit in spleen (CFU-S) and colony-forming unit in culture (CFU-C) assays were performed. Day 12 CFU-S counts of 12-week-old W/BF1 mice significantly increased in comparison with those of four-week-old W/BF1 mice or normal mice. In the CFU-C assay, CFU-GEMM and CFU-GM counts in 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or control mice. The bone marrow cells (BMCs) from 12-week-old W/BF1 mice showed a high level of G-CSF and a low level of GM-CSF in mRNA expression. To examine the effect of HSCs from 12-week-old W/BF1 mice on the onset of autoimmune diseases and the abnormal hematopoiesis, T- and B-cell-depleted BMCs of four-week-old or 12-week-old W/BF1 mice were transplanted to C3H mice. Recipient C3H mice that had received the BMCs from 12-week-old W/BF1 mice showed an earlier onset of autoimmune diseases and a shorter survival rate than those that had received the BMCs from four-week-old W/BF1 mice. These data suggest that the HSCs from 12-week-old W/BF1 mice showing the symptoms of autoimmune diseases have the capacity to induce autoimmune diseases earlier than the HSCs from four-week-old W/BF1 mice.     

Effect of chorionic gonadotrophin on hematopoietic stem cells

Department of Biological Chemistry, Perm Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR N. V. Vasil'ev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 109, No. 1, pp. 62–64, January, 1990.