Rosmarinic Acid Protects Mice from Concanavalin A-Induced Hepatic Injury through AMPK Signaling.

Rosmarinic acid (RA) is extensively utilized in herbal medicine in China. The AMP-activated protein kinase (AMPK) signaling can be activated by RA and inhibited by the synthetic, reversible AMP-competitive inhibitor, Compound C (CC). The objective of this study was to investigate the role of AMPK signaling involving the protective effects of RA on concanavalin A (Con A)-induced autoimmune hepatitis (AIH) in mice. BALB/c mice were treated with RA, with or without CC, followed by the pretreatment with Con A. Analysis of serum aminotransferases and cytokines were conducted and liver tissue histology was performed to evaluate hepatic injury. Cytokine levels in serum and hepatic tissue were respectively measured by enzyme-linked immunoassay (ELISA) and used quantitative (q)PCR. Levels of phosphorylated acetyl CoA carboxylase in the liver, representing AMPK activation, were detected by Western blotting. Compared with the Con A group, serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in RA group (100 and 150 mg/kg/d) were significantly reduced. RA also reduced hepatocyte swelling, cell death, and infiltration of leukocytes in the liver of Con A-treated mice. Serum levels of cytokines, such as interferon-γ (IFN-γ), interleukin-2 (IL-2) and interleukin-1ß (IL-1ß), were reduced by RA pretreatment, while the levels of serum interleukin-10 (IL-10), an anti-inflammatory cytokine, was elevated. These protective effects were reversed by treatment with CC. RA treatment reduced the hepatic damage via the activation of AMPK in the mice of Con A-induced. So RA acts as a potential part in the therapy of autoimmune hepatitis.

Glycyrrhizic acid ammonium salt alleviates Concanavalin A-induced immunological liver injury in mice through the regulation of the balance of immune cells and the inhibition of hepatocyte apoptosis.

Glycyrrhizic acid ammonium salt (GAAS) is derived from glycyrrhizic acid, which is an active compound extracted from the Chinese traditional medicine licorice. GAAS is clinically applied to treat immune-mediated liver injury, but its mechanism remains elusive. Therefore, this study aimed to investigate the mechanism in which GAAS alleviates immune-mediated liver injury induced by Concanavalin A (ConA). After ten days of intragastric administration of GAAS, 20 mg/kg ConA was injected via tail vein to establish the immune-mediated liver injury model of BALB/C mice. Then, the concentrations of ALT, AST, and TBIL in the serum of mice were determined. H&E staining was performed to observe the pathological changes in the liver, and the expression of liver cytokines was detected by qPCR. Immunohistochemistry and Western blot analysis was employed to detect the expression of liver-related proteins. The apoptosis in liver tissue was detected by TUNEL. Our results suggest that GAAS demonstrated excellent protective effects in the liver. We found that GAAS down-regulated the mRNA expression of IL-1ß, IL-6, TNF-α, IFN-γ, and IL-17A, and it up-regulated the mRNA expression of IL-4 and TGF-ß. Additionally, GAAS may modulate the balance of four immune cells (Th1, Th2, Th17, and Treg) by regulating the expression of T-bet, GATA3, RORγt, and Foxp3 to alleviate liver injury in mice. Furthermore, GAAS decreased hepatocyte apoptosis by blocking the JAK1/STAT1/IRF1 pathway, suppressing oxidative stress, decreasing p-JNK expression, and regulating the expression of apoptosis-related proteins. In summary, the mechanism of GAAS in liver injury alleviation acts to regulate the balance of Th cells in the liver to inhibit hepatocyte apoptosis. This study may provide a new strategy for the treatment of immune-mediated liver injury.

IgE, IgG1, and IgG4 Reactivity to Dermatophagoides pteronyssinus Glycosylated Extract in Allergic Patients.

BACKGROUND: House dust mites are important allergen sources and some of these allergenic proteins may contain carbohydrate moieties, which are able to be isolated using lectins, as Concanavalin A (ConA). This study aimed to investigate allergenicity (IgE) and antigenicity (IgG1 and IgG4) of ConA-unbound and ConA-bound Dermatophagoides pteronyssinus (Dpt) crude extracts using sera of mite-allergic patients as well as inhibition capacity of antibody binding. MATERIAL AND METHODS: We obtained mannose-enriched and mannose-depleted fractions from Dpt by ConA affinity chromatography. Both ConA-bound and ConA-unbound fractions were evaluated by ELISA and Western Blotting for specific IgE, IgG1, and IgG4 reactivity with sera obtained from 95 mite-allergic patients (DP+) and 92 nonallergic (NA) subjects. Inhibition ELISA was used to assess cross-reactivity between Dpt extract and its fractions. RESULTS: Among the DP+ patients, no difference was found between ConA-unbound and ConA-bound fractions regarding the levels of specific IgE, IgG1, and IgG4. Nonallergic subjects had the same levels of specific IgG1 to both ConA-unbound and ConA-bound fractions, although for specific IgG4, values were higher for ConA-bound. A positive correlation was found among specific IgE, IgG1, and IgG4 levels when Dpt was compared to ConA-unbound and ConA-bound fractions. Recognition of crude Dpt by IgE, IgG1, and IgG4 was highly inhibited by ConA-unbound and ConA-bound fractions. Western Blotting revealed a broad spectrum of bands ranging from 14 to 116 kDa recognized by specific IgE and IgG4. However, IgG1 reached higher frequency values on high molecular weight polypeptides. CONCLUSION: ConA-unbound and ConA-bound fractions derived from D. pteronyssinus crude extract revealed important components involved in the IgE recognition in allergic patients as well as IgG1 and/or IgG4 in allergic and healthy subjects.

Mitogen-Induced Interferon Gamma Production in Human Whole Blood: The Effect of Heat and Cations.

BACKGROUND: Interferon-gamma release assays (IGRAs) are blood tests used to measure the amount of interferon-γ (IFN-γ) released by T lymphocytes after stimulation by antigens specific for the diagnosis of latent tuberculosis infection. A mitogen serves as a positive control to assess the immune function in IGRAs. METHODS: This in vitro study was conducted to evaluate IFN-γ production by human whole blood stimulated with heat-treated and/or cation-supplemented phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM), using QuantiFERON-TB Gold Kit ELISA tests. RESULTS: The optimal concentrations of PWM, Con A and PHA for IGRAs were 2 µg/mL, 5 µg/mL and 10 µg/mL, respectively. The results showed that IFN-γ production in response to PWM was the highest and PHA was the lowest amount. The median values of three mitogens were in the following order: PWM≥Con A≥ positive control>>PHA-P>>negative control. PWM and PHA were heat stable, while Con A was heat sensitive. The mitogen response of lymphocytes to untreated or heat-treated PWM and heat-treated Con A was increased in 1 mM Ca2+-supplemented groups, whereas the response to heat-treated PHA was decreased. Exposure to 1 mM Mg2+ had no effect on untreated or heat-treated PWM, and a concentration of 1 mM Zn2+ inhibited the stimulation of un-treated PWM. We found that calcium supplementation improved the PWM-induced production of IFN-γ. CONCLUSION: Therefore, PWM is an appropriate mitogen for use as a positive control in IGRAs. It is a potential indicator of cytokine production in the diagnostic as well as research settings, and calcium supplementation improved stimulation.

Protective effects of Punica granatum (pomegranate) peel extract on concanavalin A-induced autoimmune hepatitis in mice.

Autoimmune hepatitis (AIH) is a chronic inflammatory liver disease of an unknown etiology, glucocorticoid therapy is currently recognized as an effective treatment for AIH, but conventional application and patient compliance are both hindered by its side effects. The exploration of the AIH pathogenesis and the searching for the new candidate drugs that exert potential activity and low toxicity are urgently needed. Pomegranate peel extract (PoPx) is a natural extract of Punica granatum and has been reported to have anti-inflammatory and antioxidative properties. The present study aimed to clarify the effect of PoPx on the concanavalin A (ConA)-induced autoimmune hepatitis in a mouse model that is well established at 12h after tail vein injection with a dose of 20 mg/kg of ConA. C57BL/6 female mice were pretreated with PoPx (250 mg/kg, once daily for 3 days) followed by a ConA challenge. Pretreatment with PoPx significantly alleviated ConA-induced liver injury by down-regulating the levels of plasma alanine transaminase (ALT), aspartate transaminase (AST) and cytokine, including TNF-α, interferon (IFN) -γ and interleukin (IL)-6. Moreover, liver hematoxylin and eosin (H&E) staining displayed a lighter inflammatory infiltration around the portal area in the PoPx-pretreated mice. In addition, the flow cytometry (FCM) data showed that the immune response in the liver was died down in the PoPx-pretreated condition. Specially, pretreatment with PoPx reduced the infiltration of activated CD4+ and CD8+ T cells in the liver. Taken together, these findings contributed to a better understanding of the actions of PoPx against acute AIH and indicated that PoPx might be a potential compound in treating T cell-mediated autoimmune liver injury.

Betulin from Hedyotis hedyotidea ameliorates concanavalin A-induced and T cell-mediated autoimmune hepatitis in mice.

Hedyotis hedyotidea has been used in traditional Chinese medicine for the treatment of autoimmune diseases. However, the mechanisms underlying for the effect remain unknown. We previously showed that, among 11 compounds extracted from H hedyotidea, betulin produced the strongest suppressive effect on T cell activation. Here, we examined the hepatoprotective effects of betulin against acute autoimmune hepatitis in mice and the mechanisms underlying the effects. Freshly isolated mouse splenocytes were stimulated with concanavalin A (Con A, 5 µg/mL) in the presence of betulin, the cell proliferation was assessed with CSFE-dilution assay. Mice were injected with betulin (10, 20 mg·kg-1·d-1, ip) for 3 d. One hour after the last injection, the mice were injected with Con A (15 mg/kg, iv) to induce acute hepatitis. Blood samples and liver tissues were harvested at 10 h after Con A injection, and serum transaminase levels and liver histopathology were detected; serum levels of proinflammatory cytokines, hepatic T lymphocyte ratios, and functional statuses of conventional T and NKT cells were also analyzed. Betulin (16 and 32 µmol/L) dose-dependently suppressed the proliferation of Con A-stimulated mouse splenocytes in vitro. In Con A-challenged mice, preinjection with betulin (20 mg·kg-1·d-1) significantly decreased the levels of proinflammatory cytokines IFN-γ, TNF-α and IL-6, and ameliorated liver injury. Furthermore, pretreatment with betulin (20 mg·kg-1·d-1) significantly inhibited the Con A-induced activation of NKT and conventional T cells, and decreased production of proinflammatory cytokines IFN-γ, TNF-α and IL-6 in these two cell populations. Betulin has immunomodulatory effect on overly activated conventional T and NKT cells and exerts hepatoprotective action in mouse autoimmune hepatitis. The findings provide evidence for the use of H hedyotidea and its constituent betulin in the treatment of autoimmune diseases.

Parthenolide ameliorates Concanavalin A-induced acute hepatitis in mice and modulates the macrophages to an anti-inflammatory state.

Parthenolide, the principal sesquiterpene lactone present in medicinal plants such as feverfew, has anti-microbial, anti-inflammatory and anticancer activities. In the present study, we investigated the protective role of parthenolide against acute hepatitis in mice. Mice acute hepatitis were induced by Concanavalin A and treated by parthenolide in vivo. Results shown that parthenolide remarkably reduced the congestion and necroinflammation of the mice livers with Concanavalin A-induced acute hepatitis. Meanwhile, parthenolide treatment recover the liver function which indicated by decreased the serum alanine transaminase and alkaline phosphatase activities and promoted the expression of Ki67 in the livers of these mice. In addition, parthenolide administration suppressed the Concanavalin A-induced immune reaction, as indicated by the number of F4/80, CD49b and CD4 cells present in the liver. Furthermore, parthenolide also significantly reduced the expression of pro-inflammatory cytokines such as IFN-γ, TNF-α, IL-17A, IL-1ß and IL-6 in lipopolysaccharide (LPS)-stimulated RAW264.7 cells in vitro. Moreover, parthenolide exposure decreased the phosphorylation of STAT3 and p38, and promoted the phosphorylation of p53 in RAW264.7 cells in vitro. In conclusion, parthenolide represents a drug candidate to protect the liver against Concanavalin A-induced acute hepatitis. The possible molecular mechanism involves the anti-inflammatory effects of parthenolide may by suppressing the STAT3/p38 signals and enhanced the p53 signals.

A study of the ability of bioactive extracts from brewers' spent grain to enhance the antioxidant and immunomodulatory potential of food formulations following in vitro digestion.

Bioactivity of a snack-bar, chocolate-drink and yogurt fortified with brewers' spent grain (BSG) phenolic extracts (P2 or B2) or protein hydrolysates (barley protein hydrolysate (BPH), BPH < 3 kDa, BPH < 5 kDa, BPH > 5 kDa) was measured following gastrointestinal in vitro digestion. Concentrations of 0.5 and 0.1% (v/v) digestates were chosen for addition to Caco-2 and Jurkat T cells, respectively. Yogurt and B2 digestate protected against H2O2-induced DNA damage in Caco-2 cells (p < 0.05), by the comet assay. Snack-bar digestates possessed significant (p < 0.05) immunomodulatory effects, measured by ELISA in concanavalin-A stimulated Jurkat T cells. Addition of BPH enhanced (p < 0.05) the IFN-γ reducing capacity of the snack-bar while addition of BPH < 3 and < 5 kDa reduced IL-2 production to a greater extent than unfortified yogurt (p < 0.05). Selected BSG components can enhance the antioxidant and anti-inflammatory potential of foods.

Humulus scandens exhibits immunosuppressive effects in vitro and in vivo by suppressing CD4(+) T cell activation.

Humulus scandens, rich in flavonoids, is a traditional Chinese medicine. It is widely used in China to treat tuberculosis, dysentery and chronic colitis. In this study, the major active faction of Humulus scandens (H.S) was prepared. Then, its immunosuppressive effects and underlying mechanisms on T cell activation were investigated in vitro and in vivo. Results showed that H.S significantly inhibited the proliferation of splenocytes induced by concanavalin A, lipopolysaccharides, and mixed-lymphocyte reaction in vitro. Additionally, H.S could dramatically suppress the proliferation and interferon-γ (IFN-γ) production from T cells stimulated by anti-CD3 and anti-CD28. Flow cytometric results confirmed that H.S could suppress the differentiation of IFN-γ-producing type 1 helper T cells (Th1). Furthermore, using ovalbumin immunization-induced T cell reaction and CD4(+) T-cell-mediated delayed type hypersensitivity reaction, H.S the immunosuppressive effects of H.S was also demonstrated in vivo. Western blot results showed that H.S could impede the activation of both Erk1/2 and P38 in primary T cells triggered by anti-CD3/28. Collectively, the active fraction of H.S showed promising immunosuppressive activities both in vitro and in vivo.

Rapeseed oil and ginseng saponins work synergistically to enhance Th1 and Th2 immune responses induced by the foot-and-mouth disease vaccine.

Previous investigations demonstrated that saponins isolated from the root of Panax ginseng C. A. Meyer (i.e., ginseng root saponin [GS-R]) had adjuvant activity. In the present study, the combined effects of rapeseed oil (RO) and GS-R on the immune responses elicited by foot-and-mouth disease (FMD) vaccine were investigated by measuring FMD virus (FMDV)-specific antibody levels, cytokine levels, lymphocyte proliferation, and long-lived IgG-secreting plasma cells from bone marrow in a mouse model. The results indicated that RO in combination with GS-R significantly enhanced serum IgG and isotype concentrations, gamma interferon (IFN-γ) and interleukin 5 (IL-5) levels, splenocyte proliferative responses to stimulations with concanavalin A (ConA), lipopolysaccharide (LPS), and FMDV antigen, and the numbers of IgG-secreting plasma cells in the bone marrow, suggesting that RO/GS-R enhanced both Th1 and Th2 immune responses. In addition, no significant difference was found between RO/GS-R and the commercial adjuvant oil ISA 206 in the promotion of FMD vaccine-induced immune responses. Considering the vegetable origin of RO and GS-R and the potent adjuvant activity, RO/GS-R should be studied further for the development of veterinary vaccines, especially for use in food animals in order to promote food safety.

Anti-inflammatory properties of potato glycoalkaloids in stimulated Jurkat and Raw 264.7 mouse macrophages.

AIMS: The potato glycoalkaloids, α-chaconine, α-solanine and solanidine, along with potato peel extracts were investigated for potential anti-inflammatory effects in vitro. Their potential to reduce two biomarkers of inflammation, cytokine and nitric oxide (NO) productions, were assessed in the stimulated Jurkat and macrophage models, respectively. MAIN METHODS: Cytokine and nitric oxide productions were stimulated in Jurkat and Raw 264.7 macrophages with Concanavalin A (Con A; 25 µg/ml) and lipopolysaccaride (LPS; 1 µg/ml), respectively. Selective concentrations of glycoalkaloids and potato peel extracts were added simultaneously with Con A or LPS for 24h to investigate their potential to reduce inflammatory activity. KEY FINDINGS: α-Chaconine and solanidine significantly reduced interleukin-2 (IL-2) and interleukin-8 (IL-8) productions in Con A-induced Jurkat cells. The potato peel extracts did not influence cytokine production. In LPS-stimulated Raw macrophages, α-solanine, solanidine and two potato peel extracts significantly reduced induced NO production. SIGNIFICANCE: Our findings suggest that sub-cytotoxic concentrations of potato glycoalkaloids and potato peel extracts possess anti-inflammatory effects in vitro and with further investigation may be useful in the prevention of anti-inflammatory diseases.

Influence of the slow infusion of a soybean oil emulsion on plasma cytokines and ex vivo T cell proliferation after an esophagectomy.

BACKGROUND: Lipid emulsions have been suggested to reduce immune responses, particularly in severely stressed patients. The authors investigated the influence of the slow intravenous infusion of a soybean oil-based lipid emulsion on some immune parameters in patients who had undergone an esophagectomy for esophageal cancer. METHODS: Thirty-two patients who had undergone an esophagectomy were randomly divided into a lipid emulsion (LPD)-treated group and a control group. All patients received parenteral feeding with a glucose-based solution. Patients in the LPD group received 100 mL of a 20% soybean oil emulsion for 7 days after the esophagectomy in addition to the glucose-based feeding. A slow infusion rate (0.09-0.12 g/kg/h) was adopted to take account of the intrinsic degradation of infused lipids. Immune responses were measured based on lymphocyte proliferation and serum concentrations of monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). The authors also measured levels of rapid turnover proteins (ie, transferrin, prealbumin, and retinol-binding protein). RESULTS: Phytohemagglutinin- and concanavalin A-stimulated lymphocyte proliferation significantly decreased after the esophagectomy, but no significant difference was seen between the LPD and control groups. No significant difference in changes in plasma concentrations of MCP-1, IL-6 and TNF-α occurred between the 2 groups either. Plasma concentrations of rapid turnover proteins did not differ between the groups. CONCLUSIONS: These results indicate that the lipid emulsion did not affect the immune parameters measured in patients who had undergone an esophagectomy when administered at a slow rate.

Immune response and blood chemistry of pigs fed conjugated linoleic acid.

Immune function (response to concanavalin A, cytokine production, and lymphocyte profiles) and blood chemistry variables were measured in growing-finishing pigs (Yorkshire/Landrace/Duroc dam × Hampshire sire) fed varying percentages of CLA (0, 0.12, 0.25, 0.50, and 1.0%). Blood was collected at 0, 14, 28, 42, and 56 d on feed (DOF). Total white blood cell (WBC) count increased (P < 0.01) linearly to 42 DOF. No differences (P = 0.53) were observed for WBC across CLA treatment. Nitric oxide was greater (P < 0.01) for the 1.0% CLA treatment compared with all other treatments. Flow cytometry using fluorescent labeled monoclonal antibodies to the CD4, CD8, double-positive CD4/CD8, and CD2 surface markers was used to determine lymphocyte subpopulations. Supplementation of CLA had no effect (P = 0.61) on lymphocyte subpopulation cell distribution. Most blood chemistry variables were within the normal metabolic range for pigs. A decrease was observed over DOF for P (P < 0.01) and K (P < 0.05). Additionally, Na and Cl concentrations increased (P < 0.05) from 14 to 28 DOF and decreased over the remainder of the trial. Electrolyte balance was not different (P = 0.38) across CLA treatments and was likely explained by no differences in feed intake among the CLA treatment groups. Blood lipid variables indicated that total cholesterol (P < 0.001), triglycerides (P < 0.001), high-density lipoproteins (P < 0.001), and low-density lipoproteins (P < 0.01) increased as the amount of CLA in the diet increased, but none of the results from these treatments exceeded the normal range of acceptability. These results suggested that CLA was safe when fed to growing-finishing pigs and had little effect on their immune function and blood chemistry variables.

Immunosuppressive activity on the murine immune responses of glycyrol from Glycyrrhiza uralensis via inhibition of calcineurin activity.

CONTEXT: Calcineurin (CN), a unique protein phosphatase, plays an important role in immune regulation. Our laboratory has established an effective molecular drug-screening model based on CN activity. OBJECTIVE: Our aim is to search for an effective immunosuppressant from Glycyrrhiza uralensis (Leguminosae). MATERIALS AND METHODS: As guided by CN inhibitory test, an active compound was purified and identified as glycyrol. Immunosuppressive activity of glycyrol in vitro was assayed by T lymphocytes proliferation and mixed lymphocyte reaction (MLR). In addition, delayed-type hypersensitivity reaction (DTH) and skin allograft test in vivo were also carried out. Further, we have investigated the effect of glycyrol on phorbol 12-myristate 13-acetate (PMA)/ionomycin (Io)-stimulated IL-2 expression in Jurkat cells. RESULTS: The enzymatic assay showed glycyrol (IC(50) = 84.6 µM) inhibited calcineurin activity in a dose-dependent manner. Glycyrol, at the non-cytotoxic concentration, significantly inhibited proliferation of murine spleen T lymphocytes induced by Concanavalin A (Con A) and mixed lymphocyte reaction (MLR) in vitro. In addition, mice treated with glycyrol had shown the dose-dependent decrease in delayed type hypersensitivity (DTH) and prolonged the graft survival by 59% compared to the control group (*p < 0.05). RT-PCR showed glycyrol suppressed IL-2 production in a concentration-dependent manner. DISCUSSION AND CONCLUSION: Our results show the immunosuppressive activity of glycyrol and this activity should be due to its inhibitory effect on CN activity, thereby suppressing IL-2 production and regulating T lymphocytes. Thus, glycyrol could be a candidate for development as a novel immunomodulatory drug.

Effect of L-carnitine on proliferative response and mRNA expression of some of its associated factors in splenic mononuclear cells of male broiler chicks.

The effect of L-carnitine supplementation on mitogen (concanavalin A, Con A) induced proliferation of mononuclear cells (MNC) in the spleen was investigated in broiler chickens at different ages. Day-old chickens were fed a diet supplemented with or without L-carnitine (100 ppm) for 24 days. The carnitine-supplemented group showed greater proliferation of MNC in the spleen in response to Con A than that of the control group at 24 days of age. In addition, at 24 days of age the carnitine-supplemented group showed higher expression of interleukin (IL)-2 and interferon (IFN)-gamma mRNA, but lower expression of inducible nitric oxide synthase (iNOS) in the Con A-stimulated splenic MNC than the control group. The enhancement effect of L-carnitine on MNC proliferation and IL-2 mRNA expression was not found in chicks at 14 days of age. Addition of L-carnitine (50 nmol/mL) to the culture medium enhanced proliferation and IL-2 mRNA expression of splenic MNC obtained from 24-day-old but not from 14-day-old broiler chickens. The results suggest that L-carnitine is capable of enhancing MNC proliferation in broiler chickens at 24 days of age partly through increasing IL-2 and IFN-gamma production and decreasing NO production.

Changes of color and blood flow of the tongue in the mini-swine of immune hepatic injury.

OBJECTIVE: To investigate color and microvascular blood flow of the tongue in the mini-swine with immune hepatic injury. METHODS: Six Chinese mini-swine for experimental use, 3 males and 3 females, were randomly divided into two groups, normal group and model group, 3 swine in each group. The swine in the model group was administrated by injection of 5 mg/kg ConA into the vein of auricular back, once every other day, 3 times each week, for 2 weeks in total. The animal in the control group was administrated with equal volume of saline. At 9 o'clock in the morning of the 15th day of the experiment, each swine was anesthetized with intramuscular injection of 9 ml 2.5% pentobarbital sodium and 3 ml Maleate, and then picture of the tongue was taken, microvascular blood flow on the tongue and the liver was detected with a laser Doppler blood flowmeter; Blood was taken from the precaval vein. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (Tbil) and total protein (TP) were determined; Pathological changes of the liver and tongue tissues were investigated by means of HE staining; Serum TNF-alpha content was detected with ELISA assay. RESULTS: In the mini-swine with immune hepatic injury induced by ConA, the tongue color showed cyanotic color, microvascular perfusion in the liver and the tongue, and partial pressure of oxygen in the tongue tissue significantly decreased; and the microcirculatory perfusion of the tongue was significantly correlated with that of the liver and the HIS color spatial value of the tongue; Serum TNF-alpha content significantly increased. CONCLUSION: The mini-swine with immune hepatic injury induced by ConA conforms to pathological characteristics of immune hepatic injury. Formation of the cyanotic tongue is related with microcirculatory disturbance of the tongue, which can indirectly reflect hepatic microcirculatory state in the immune hepatic injury.

Ganoderma lucidum extracts inhibited leukemia WEHI-3 cells in BALB/c mice and promoted an immune response in vivo.

Ganoderma lucidum (G. lucidum) is a medicinal mushroom having biological effects such as immunomodulation and anti-tumor actions. In China and many other Asian countries, G. lucidum is used as a folk remedy to promote health and longevity. Although many studies have shown that G. lucidum modulates the immune system, including, for example, antigen-presenting cells, natural killer (NK) cells, and the T and B lymphocytes, the effects of G. lucidum on the WEHI-3 leukemic BALB/c mice are unclear. We attempted to determine whether G. lucidum would promote immune responses in BALB/c mice injected with WEHI-3 leukemia cells. The effects of G. lucidum on the survival rate of WEHI-3 leukemia cells injected into BALB/c mice were examined. It increased the percentages of CD3 and CD19, but decreased the percentages of Mac-3 and CD11b markers, suggesting that differentiation of the precursor of T and B cells was promoted but macrophages were inhibited. It decreased the weight of spleens as compared with control mice. It also promoted phagocytosis by macrophage from peripheral blood mononuclear cell (PBMC) and it also promoted natural killer cell activity. It decreased the percentage of leukemia cells in the spleens of mice before they were injected with WEHI-3 cells. Apparently, G. lucidum affects murine leukemia WEHI-3 cells in vivo.

Improvement of leucocyte functions in ovariectomised aged rats after treatment with growth hormone, melatonin, oestrogens or phyto-oestrogens.

Ageing is accompanied by an impairment of the physiological activity of the nervous, endocrine and immune system, as well as in neuroendocrine-immune communication. However, age-related changes in this communication axis have been scarcely studied. In mammals, the process of ageing is associated with an important decline in the secretion of several hormones, such as growth hormone (GH), melatonin (MEL) and oestrogens (Os). Ovariectomy, a model of menopause in rats, has been found to lead to premature immunosenescence. In the present study, the effect of ovariectomy and the role of replacement therapies with GH, MEL, O and natural phyto-oestrogens (POs) have been assessed on several functions in leucocytes from the spleen and the axillary nodes of intact and ovariectomised rats. Chemotaxis, lymphoproliferative response to the mitogen concanavalin A (Con A), the release of interleukin-2 (IL-2) and the natural killer (NK) cell activity have been investigated. Age-controlled rats were used to compare immune functions in hormone treated aged rats with those in younger untreated animals. In all experimental groups, the immune impairment caused by ageing and ovariectomy was partially or completely reversed by hormone treatments. Since the immune system is a marker of health and a predictor of longevity, the results suggest that treatment with hormones could slow down the effects of the ageing process.

Scrophularia buergeriana regulates cytokine production in vitro.

The Scrophularia buergeriana (SB) has long been used to treat various diseases an account of its antimicrobial and anti-virus activity. However, it is unclear how SB regulates the immune responses. This study investigated the effect of SB on the production of cytokines in a human T-cell line, MOLT-4 cells, and mouse peritoneal macrophages. The MOLT-4 cells were cultured for 24 h in the presence or absence of SB plus concanavalin (con) A. SB plus con A significantly increased the level of interleukin (IL)-2, IL-4 and interferon (IFN)-gamma production compared with that of con A alone (approximately 1.79-fold for IL-2, 2-fold for IL-4, and 1.85-fold for IFN-gamma, p < 0.05). SB plus recombinant IFN-gamma (rIFN-gamma) increased the level of IL-12 and NO production compared with rIFN-gamma alone. In addition, SB plus rIFN-gamma increased the level the iNOS expression on mouse peritoneal macrophages. Overall, SB may have an immune-enhancement effect through cytokine production.

[Effect of ronggan mixture on cell apoptosis in rats with chronic immune liver injury induced by concanavalin A].

OBJECTIVE: To explore the effects of Ronggan Mixture (RGM) on cell apoptosis by observing the expressions of apoptosis-related genes (Fasl and Bcl-2) in transgenic mice with chronic liver immune injury induced by concanavalin A (ConA). METHODS: Seventy-four transgenic mice were divided into 6 groups, the model group, the normal group, and the treated groups treated respectively with biphenyldicarboxylate (DDB), oriental wormwood (OWW), Yinchenhao Decoction (YCHD) and RGM. Pathologic changes of liver tissue were observed by light microscopy, number of apoptotic cells were determined by TUNEL method, and expressions of apoptosis-related genes, Fasl and Bcl-2, in hepatic T lymphocyte were detected by flow cytometer. RESULTS: Evident pathological changes of liver appeared in the model mice, showed severely destroyed structure of hepatic lobules. As compared with the model group, the changes of liver fibrosis and cell necrosis were much lessened in the RGM group and the YCHD group (P < 0.05). The protein expression of apoptotic gene Fasl and the apoptotic index in the model group were higher than those in the normal group (P < 0.05), but that of the apoptotic inhibiting gene, Bcl-2, in model mice was similar to that in normal mice. As compared with the model group, apoptosis index decreased (P < 0.01), levels of Fasl expression was lower and Bcl-2 expression was higher in the RGM group and the YCHD group (P < 0.05, P < 0.01), and the effect of the two was similar, but significantly superior to that of OWW and DDB (P < 0.05 or P < 0.01). CONCLUSION: The Chinese compound, RGM and YCHD can not only relieve the hepatic pathological injury, but also reduce the cell apoptosis in chronic liver immune injury mice through regulating the expressions of Fasl and Bcl-2.