BaZiBuShen alleviates cognitive deficits and regulates Sirt6/NRF2/HO-1 and Sirt6/P53-PGC-1α-TERT signaling pathways in aging mice.

ETHNOPHARMACOLOGICAL RELEVANCE: BaZiBuShen formula (BZBS) is clinically used to counteract mental fatigue and to retard the aging process. Brain aging echoes in major risks of human sufferings and has become one of the main challenges to our societies and the health-care systems. AIM OF THE STUDY: To investigate the effect and mode of action of BZBS on aging-associated cognitive impairments. MATERIALS AND METHODS: BZBS was orally administered to D-galactose and NaNO2-induced aging mice. Premature senescence was assessed using the Morris water maze, step-down type passive avoidance, and pole-climbing tests. Telomere length was examined by qPCR analysis. Telomerase activity was assessed using PCR ELISA assay. Mitochondrial complex IV activity was examined by biochemical test. The levels of redox and immune status were determined by ELISA or biochemical assay. The expressions of sirtuin 6 (Sirt6), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), P53, telomerase reverse transcriptase (TERT), heme oxygenase-1 (HO-1), phospho(p)-nuclear factor erythroid-2 related factor 2 (NRF2), caspase-3, Bcl-2 associated x (Bax), and B-cell lymphoma-2 (Bcl-2) in the cerebral cortex were examined by Western blot and/or immunohistochemical staining. RESULTS: BZBS intervention ameliorated reduced brain performances in aging mice, including memory, cognitive, and motor functions. In addition, BZBS administration to aging mice preserved redox homeostasis, attenuated immunosenescence, and maintained telomerase activity and telomere length. Moreover, BZBS treatment were associated with a declines in P53, caspase-3, Bax expressions and an increase in Sirt6, p-HO-1, p-NRF2, PGC-1α, and Bcl-2 expressions in the brains of this rapid aging mouse. CONCLUSIONS: BZBS attenuates premature senescence possibly via the preservation of redox homeostasis and telomere integrity, and inhibition of apoptosis in rapid aging mouse. The mechanism governing the alterations may be associated with through the activation of Sirt6/NRF2/HO-1 and Sirt6/P53-PGC-1α-TERT signaling pathways. The results suggest that BZBS may provide a novel strategy for confronting aging and age-associated diseases.

Protocatechuic Acid Inhibits Vulnerable Atherosclerotic Lesion Progression in Older Apoe-/- Mice.

BACKGROUND: Normalization of arterial inflammation inhibits atherosclerosis. The preventive role for protocatechuic acid (PCA) in early-stage atherosclerosis is well recognized; however, its therapeutic role in late-stage atherosclerosis remains unexplored. OBJECTIVE: We investigated whether PCA inhibits vulnerable atherosclerosis progression by normalizing arterial inflammation. METHODS: Thirty-wk-old male apolipoprotein E-deficient (Apoe-/-) mice with vulnerable atherosclerotic lesions in the brachiocephalic artery were fed the AIN-93G diet alone (control) or supplemented with 0.003% PCA (wt:wt) for 20 wk. Lesion size and composition, IL-1ß, and NF-κB in the brachiocephalic arteries, and serum lipid profiles, oxidative status, and proinflammatory cytokines (e.g., IL-1ß, monocyte chemoattractant protein-1, and serum amyloid A) were measured. Moreover, the effect of PCA on the inflammation response was evaluated in efferocytic macrophages from C57BL/6J mice. RESULTS: Compared with the control treatment, dietary PCA supplementation significantly reduced lesion size (27.5%; P < 0.05) and also improved lesion stability (P < 0.05) as evidenced by increased thin fibrous cap thickness (31.7%) and collagen accumulation (58.3%), reduced necrotic core size (37.6%) and cellular apoptosis (73.9%), reduced macrophage accumulation (45.1%), and increased vascular smooth muscle cell accumulation (51.5%). Moreover, PCA supplementation inhibited IL-1ß expression (53.7%) and NF-κB activation (64.4%) in lesions. However, PCA supplementation did not change serum lipid profiles, total antioxidant capacity, and inflammatory cytokines. In efferocytic macrophages, PCA at 0.5 and 1 µmol/L inhibited Il1b/IL-1ß mRNA (27.2-46.5%) and protein (29.2-49.6%) expression and NF-κB activation (67.0-80.3%) by upregulation of MER proto-oncogene tyrosine kinase (MERTK) and inhibition of mitogen-activated protein kinase 3/1 (MAPK3/1). Strikingly, the similar pattern of the MERTK and MAPK3/1 changes in lesional macrophages of mice after PCA intervention in vivo was recapitulated. CONCLUSION: PCA inhibits vulnerable lesion progression in mice, which might partially be caused by normalization of arterial inflammation by upregulation of MERTK and inhibition of MAPK3/1 in lesional macrophages.

Suppression of miR-21 and miR-155 of macrophage by cinnamaldehyde ameliorates ulcerative colitis.

Ulcerative colitis (UC) is a major form of inflammatory bowel disease which involved mucosal immune dysfunction. Cinnamaldehyde (CA) is major active compound from cinnamon, a useful traditional medicine in Asia which shows superior antibacterial and anti-inflammatory activity. In this study, we investigated the effects of CA on UC both in vivo and in vitro. We showed that CA attenuated the symptoms of DSS-induced colitis, including loss of body weights, disease activity index (DAI), shortening of the colon lengths and infiltration of inflammatory cells. Moreover, CA decreased the pro-inflammatory cytokines and NLRP3 inflammasome, miR-21 and miR-155 in colon tissues, in addition, the percentage of macrophages was reduced based on the surface marker F4/80 and IL-10 secretion in CA-treated group, suggesting that the CA ameliorate the UC via activation of macrophage. Herein, the effects of CA on macrophage cells were examined in vitro. We found that CA reduced the level of proinflammatory cytokines, such as TNF-α, IL-1ß, IL-6, in the activation of RAW264.7, human macrophage-like cells U937, and primary peritoneal macrophages. Furthermore, the suppression of NLRP3 inflammasome, miR-21 and miR-155 was also found in CA-treated LPS-stimulated RAW264.7 cells. CA also reduced the production of reactive oxygen species, the phosphorylation of AKT, mTOR and COX2 protein level in the RAW264.7. Meanwhile, data revealed that transferred miR-21 or miR-155 inhibitor suppressed levels of IL-1ß and IL-6, whereas miR-21 or miR-155 mimics increased expressions of these, and CA suppressed these expressions. Our results indicate that CA could ameliorate DSS-induced colitis through inhibition of NLRP3 inflammasome activation and miR-21 and miR-155 levels in colons and macrophage, suggesting that CA might be a potentially effective drug for UC.

Liver X Receptor Agonist Therapy Prevents Diffuse Alveolar Hemorrhage in Murine Lupus by Repolarizing Macrophages.

The generation of CD138+ phagocytic macrophages with an alternative (M2) phenotype that clear apoptotic cells from tissues is defective in lupus. Liver X receptor-alpha (LXRα) is an oxysterol-regulated transcription factor that promotes reverse cholesterol transport and alternative (M2) macrophage activation. Conversely, hypoxia-inducible factor 1-α (HIF1α) promotes classical (M1) macrophage activation. The objective of this study was to see if lupus can be treated by enhancing the generation of M2-like macrophages using LXR agonists. Peritoneal macrophages from pristane-treated mice had an M1 phenotype, high HIFα-regulated phosphofructokinase and TNFα expression (quantitative PCR, flow cytometry), and low expression of the LXRα-regulated gene ATP binding cassette subfamily A member 1 (Abca1) and Il10 vs. mice treated with mineral oil, a control inflammatory oil that does not cause lupus. Glycolytic metabolism (extracellular flux assays) and Hif1a expression were higher in pristane-treated mice (M1-like) whereas oxidative metabolism and LXRα expression were higher in mineral oil-treated mice (M2-like). Similarly, lupus patients' monocytes exhibited low LXRα/ABCA1 and high HIF1α vs. CONTROLS: The LXR agonist T0901317 inhibited type I interferon and increased ABCA1 in lupus patients' monocytes and in murine peritoneal macrophages. In vivo, T0901317 induced M2-like macrophage polarization and protected mice from diffuse alveolar hemorrhage (DAH), an often fatal complication of lupus. We conclude that end-organ damage (DAH) in murine lupus can be prevented using an LXR agonist to correct a macrophage differentiation abnormality characteristic of lupus. LXR agonists also decrease inflammatory cytokine production by human lupus monocytes, suggesting that these agents may be have a role in the pharmacotherapy of lupus.

Tanshindiol C inhibits oxidized low-density lipoprotein induced macrophage foam cell formation via a peroxiredoxin 1 dependent pathway.

NF-E2-related factor 2 (Nrf2) has been shown to be protective in atherosclerosis. The loss of Nrf2 in macrophages enhances foam cell formation and promotes early atherogenesis. Tanshindiol C (Tan C) is isolated from the root of Salvia miltiorrhiza Bge., a traditional Chinese medicine that has been used for the treatment of several cardiovascular diseases for many years. This study was aimed to test the potential role of Tan C against macrophage foam cell formation and to explore the underlying mechanism. Firstly, we observed that Tan C markedly suppressed oxidized low-density lipoprotein (oxLDL) induced macrophage foam cell formation. Then, we found that Tan C was an activator of both Nrf2 and Sirtuin 1 (Sirt1) in macrophages. Nrf2 and Sirt1 synergistically activated the transcription of anti-oxidant peroxiredoxin 1 (Prdx1) after Tan C treatment. More important, we demonstrated that silencing of Prdx1 promoted oxLDL-induced macrophage foam cell formation. Prdx1 upregulated adenosine triphosphate-binding cassette (ABC) transporter A1 (ABCA1) expression and decreased intracellular lipid accumulation. Furthermore, Tan C ameliorated oxLDL induced macrophage foam cell formation in a Prdx1-dependent manner. These observations suggest that Tan C protects macrophages from oxLDL induced foam cell formation via activation of Prdx1/ABCA1 signaling and that Prdx1 may be a novel target for therapeutic intervention of atherosclerosis.

Compatibility of Tanshinone IIA and Astragaloside IV in attenuating hypoxia-induced cardiomyocytes injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Herbal medicines including Tanshinone IIA (TanIIA) and Astragaloside IV (AsIV) are widely used in Asia as therapeutic agents for cardiovascular diseases, due to their complementary roles and shared properties based on the theory of traditional Chinese medicine and pharmacological researches. However, the underlying pathological mechanisms for their efficacy are still unclear. In addition, the compatibility or incompatibility of the herbal medicines when administered with other herbal remedies or with prescription drugs is unknown. AIM OF THE STUDY: We aimed to investigate the compatibility of TanIIA and AsIV in protecting cardiomyocytes against hypoxia-induced injury. MATERIALS AND METHODS: Cultured cardiomyocytes were stimulated in hypoxia condition, in the absence or presence of the two herbal compounds, TanIIA and AsIV. Indicators were determined by cytotoxicity assay, quantitative PCR, ELISA, flow cytometry assay, immunofluorescence staining and western blot. RESULTS: Either TanIIA alone or the combined herbal compounds inhibited hypoxia-triggered chemokines production including CCL2/5/19, CXCL2 and Transwell assay-indicated monocyte/macrophage recruitment, cytokines production including TNF-α and IL-6. While AsIV alone or the combined herbal compounds attenuated hypoxia-induced cell apoptosis indicated by decreased Annexin V+ cells and the ratio of Bax/Bcl-2, but no significant effect of the herbal compounds was observed in modulating cell apoptosis following both hypoxia and TNF-α stimulation. As an anti-apoptotic factor, stress granule formation was further enhanced by AsIV alone or the combined herbal compounds in hypoxia or heat shock stress. Moreover, immunoblotting analysis indicated that stress-responsive mitogen-activated protein kinases (MAPK) pathways including the phosphorylation of ERK1/2, p38 and JNK were inhibited while the phosphorylation of Akt in phosphatidylinositol 3-kinase (PI3K) -Akt pathway for cell survival was restored by the herbal compounds. Among these results, the combination of TanIIA and AsIV comprised most of the beneficial properties tested, although their combination did not improve the maximal effects achieved by any of the compounds alone. CONCLUSIONS: Taken together, these data suggest a compatibility of TanIIA and AsIV in protecting cardiomyocyte against hypoxia-induced injury.

Supplementation with linoleic acid-rich soybean oil stimulates macrophage foam cell formation via increased oxidative stress and diacylglycerol acyltransferase1-mediated triglyceride biosynthesis.

During the last decades there has been a staggering rise in human consumption of soybean oil (SO) and its major polyunsaturated fatty acid linoleic acid (LA). The role of SO or LA in cardiovascular diseases is highly controversial, and their impact on macrophage foam cell formation, the hallmark of early atherogenesis, is unclear. To investigate the effects of high SO or LA intake on macrophage lipid metabolism and the related mechanisms of action, C57BL/6 mice were orally supplemented with increasing levels of SO-based emulsion or equivalent levels of purified LA for 1 month, followed by analyses of lipid accumulation and peroxidation in aortas, serum and in peritoneal macrophages (MPM) of the mice. Lipid peroxidation and triglyceride mass in aortas from SO or LA supplemented mice were dose-dependently and significantly increased. In MPM from SO or LA supplemented mice, lipid peroxides were significantly increased and a marked accumulation of cellular triglycerides was found in accordance with enhanced triglyceride biosynthesis rate and overexpression of diacylglycerol acyltransferase1 (DGAT1), the key enzyme in triglyceride biosynthesis. In cultured J774A.1 macrophages treated with SO or LA, triglyceride accumulated via increased oxidative stress and a p38 mitogen-activated protein kinase (MAPK)-mediated overexpression of DGAT1. Accordingly, anti-oxidants (pomegranate polyphenols), inhibition of p38 MAPK (by SB202190) or DGAT1 (by oleanolic acid), all significantly attenuated SO or LA-induced macrophage triglyceride accumulation. These findings reveal novel mechanisms by which supplementation with SO or LA stimulate macrophage foam cell formation, suggesting a pro-atherogenic role for overconsumption of SO or LA. © 2016 BioFactors, 43(1):100-116, 2017.

Baicalin ameliorates experimental inflammatory bowel disease through polarization of macrophages to an M2 phenotype.

Inflammatory bowel diseases (IBDs) are chronic inflammatory disorders of the intestinal tract. Baicalin, originally isolated from the root of the Chinese herb Huangqin (Scutellaria baicalensis Georgi) and its main active ingredient, has a protective effect against inflammatory responses in several diseases. The present study investigated the effects of baicalin on macrophage polarization and its therapeutic role in IBD. Murine peritoneal macrophages and mice with colitis were treated with baicalin. Macrophage subset distribution, M1 and M2 macrophage-associated mRNA expression, and interferon regulatory factor 4 and 5 (IRF4 and IRF5) expression were analyzed. siRNA transfection into mouse peritoneal macrophages was utilized to suppress IRF4. Fluorescence-activated cell sorting, western blot, and real-time PCR analyses were performed. Baicalin (50µM) limited lipopolysaccharide (LPS)-induced M1 macrophage polarization; decreased LPS-induced tumor necrosis factor α, interleukin (IL)-23, and IRF5 expression; and increased IL-10, arginase-1 (Arg-1), and IRF4 expression. siRNA-mediated IRF4 silencing significantly impaired baicalin activity. Furthermore, pretreatment with baicalin (100mg/kg) in mice with dextran sodium sulfate (DSS)-induced colitis ameliorated the severity of colitis and significantly decreased the disease activity index (baicalin group, 3.33±0.52 vs. DSS group, 5.67±1.03). Baicalin (100mg/kg) also repressed IRF5 protein expression and promoted IRF4 protein expression in the lamina propria mononuclear cells, and induced macrophage polarization to the M2 phenotype. In summary, our results showed that baicalin upregulates IRF4 protein expression and reverses LPS-induced macrophage subset redistribution. Thus, baicalin alleviates DSS-induced colitis by modulating macrophage polarization to the M2 phenotype.

The Impaired Function of Macrophages Induced by Strenuous Exercise Could Not Be Ameliorated by BCAA Supplementation.

The aim of this study was to evaluate the effect of strenuous exercise on the functions of peritoneal macrophages in rats and to test the hypothesis that branched-chain amino acid (BCAA) supplementation will be beneficial to the macrophages of rats from strenuous exercise. Forty male Wistar rats were randomly divided into five groups: (C) Control, E) Exercise, (E1) Exercise with one week to recover, (ES) Exercise + Supplementation and (ES1) Exercise + Supplementation with 1 week to recover. All rats except those of the sedentary control were subjected to four weeks of strenuous exercise. Blood hemoglobin, serum testosterone and BCAA levels were tested. Peritoneal macrophages functions were also determined at the same time. The data showed that hemoglobin, testosterone, BCAA levels, and body weight in group E decreased significantly as compared with that of group C. Meanwhile, phagocytosis capacity (decreased by 17.07%, p = 0.031), reactive oxygen species (ROS) production (decreased by 26%, p = 0.003) and MHC II mRNA (decreased by 22%, p = 0.041) of macrophages decreased in the strenuous exercise group as compared with group C. However, the chemotaxis of macrophages did not change significantly. In addition, BCAA supplementation could slightly increase the serum BCAA levels of rats from strenuous exercise (increased by 6.70%, p > 0.05). Moreover, the body weight, the blood hemoglobin, the serum testosterone and the function of peritoneal macrophages in group ES did not change significantly as compared with group E. These results suggest that long-term intensive exercise impairs the function of macrophages, which is essential for microbicidal capability. This may represent a novel mechanism of immunosuppression induced by strenuous exercise. Moreover, the impaired function of macrophage induced by strenuous exercise could not be ameliorated by BCAA supplementation in the dosing and timing used for this study.

Omega 3 Polyunsaturated Fatty Acids Suppress the Development of Aortic Aneurysms Through the Inhibition of Macrophage-Mediated Inflammation.

BACKGROUND: Dietary intake of ω3 polyunsaturated fatty acids (ω3-PUFAs) reduces progression of atherosclerosis and prevents future cardiovascular events. Macrophages are key players in the pathogenesis of aortic aneurysm. The effects of ω3-PUFAs on abdominal aortic aneurysm (AAA) formation and macrophage-mediated inflammation remain unclear. METHODS AND RESULTS: The AAA model was developed by angiotensin II infusion in apolipoprotein E-deficient mice. Mice were supplemented with eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA). The development of AAA lesions and macrophage infiltration in the aorta were analyzed. Gene expression of inflammatory markers in aortic tissues and peritoneal macrophages were measured by using quantitative polymerase chain reaction. AAA formation and macrophage infiltration were significantly suppressed after EPA and DHA administration. EPA administration and DHA administration significantly decreased the expression of tumor necrosis factor-α, monocyte chemoattractant protein-1, transforming growth factor-ß, matrix metalloproteinases (MMP)-2, MMP-9, and vascular cell adhesion molecule-1 in the aortas. The expression of arginase 2, which is a marker of pro-inflammatory macrophages, was significantly lower and that of Ym1, which is a marker of anti-inflammatory macrophages, and was significantly higher after EPA and DHA administration. The same trends were observed in peritoneal macrophages after EPA and DHA administration. CONCLUSIONS: Dietary intake of EPA and DHA prevented AAA development through the inhibition of aortic and macrophage-mediated inflammation.

Combination use of ferulic acid, ligustrazine and tetrahydropalmatine inhibits the growth of ectopic endometrial tissue: a multi-target therapy for endometriosis rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ferulic acid (FA), ligustrazine (LZ) and tetrahydropalmatine (THP) are separately isolated from Chinese Angelica, Szechwan Lovage Rhizome and Rhizoma in the Jiawei-Foshou-San formula, a popular traditional Chinese medicine for irregular menses. It has been reported that the combination use of FA+LZ+THP has similar effect on endometriosis, but the underlying mechanisms are unclear. This study was to investigate the combination effects and mechanisms of FA+LZ+THP on endometriosis rats. MATERIALS AND METHODS: Fifty endometriosis rats were intragastricly treated with FA+LZ+THP for 4 wk. The volume of ectopic endometrial tissue was measured to evaluate the effects. Then the changes in hypothalamic-pituitary-ovarian axis and ERE pathway were indicated by the levels of E2, GnRH, FSH and LH, and the expressions of ER, HSP90 and COX-2, respectively. In addition, peritoneal macrophages of each rat were cultured in vitro and treated with (FA+LZ+THP)-medicated serum for 24h. The proliferation and phagocytosis abilities, the levels of IL-1ß and TNF-α, and the expression of IκBα were then measured for the changes of peritoneal macrophage activities. RESULTS: Combination use of FA+LZ+THP diminished the volume of the ectopic endometrial tissues (P<0.05 or P<0.01). It also decreased the E2 level, suppressed the expression of GnRH, FSH and LH, and decreased the protein expression of ER, HSP90 and COX-2 (all P<0.05 or P<0.01). The phagocytosis ability of peritoneal macrophage was enhanced by (FA+LZ+THP)-medicated serum (P<0.05) with no change of proliferation (P>0.05). Moreover, IL-1ß and TNF-α were downregulated (both P<0.05 or P<0.01) and IκBα was upregulated by the (FA+LZ+THP)-medicated serum (P<0.01). CONCLUSIONS: The combination use of FA, LZ and THP could inhibit the growth of ectopic endometrial tissue in endometriosis rats. It might be related to the down-regulation of hypothalamic-pituitary-ovarian axis, the amelioration in ERE pathway and the improvement of peritoneal macrophage activities.

The preparation of gypenosides liposomes and its effects on the peritoneal macrophages function in vitro.

Optimization of preparation of gypenosides liposome (GPSL) was investigated with response surface methodology (RSM) and the effect of GPSL activating phagocytosis activity and cytokine secretion of macrophages was determined in the paper. The results showed that the optimal conditions of GPSL preparation were as follows: the ratio of lipid to drug (w/w) of 8:1, the ratio of soybean phospholipid to cholesterol (w/w) of 6:1, the temperature of incubation of drug in water bath of 55°C, the time of incubation of drug of 11 min. With the condition, the experimental encapsulation efficiency of GPSL was 90.17 ± 0.38%. In addition, GPSL could significantly enhance phagocytosis activity and the secretion of cytokines of the peritoneal macrophages of mice. These indicated that GPSL with high entrapping efficiency and small particle size could be prepared by ethanol injection combined with ammonium sulfate gradient method, which is applied easily, and GPSL could enhance the activity of peritoneal macrophages.

Macrophage-derived matrix vesicles: an alternative novel mechanism for microcalcification in atherosclerotic plaques.

RATIONALE: We previously showed that early calcification of atherosclerotic plaques associates with macrophage accumulation. Chronic renal disease and mineral imbalance accelerate calcification and the subsequent release of matrix vesicles (MVs), precursors of microcalcification. OBJECTIVE: We tested the hypothesis that macrophage-derived MVs contribute directly to microcalcification. METHODS AND RESULTS: Macrophages associated with regions of calcified vesicular structures in human carotid plaques (n=136 patients). In vitro, macrophages released MVs with high calcification and aggregation potential. MVs expressed exosomal markers (CD9 and TSG101) and contained S100A9 and annexin V. Silencing S100A9 in vitro and genetic deficiency in S100A9-/- mice reduced MV calcification, whereas stimulation with S100A9 increased calcification potential. Externalization of phosphatidylserine after Ca/P stimulation and interaction of S100A9 and annexin V indicated that a phosphatidylserine-annexin V-S100A9 membrane complex facilitates hydroxyapatite nucleation within the macrophage-derived MV membrane. CONCLUSIONS: Our results support the novel concept that macrophages release calcifying MVs enriched in S100A9 and annexin V, which contribute to accelerated microcalcification in chronic renal disease.

Immunomodulatory and cellular anti-oxidant activities of an aqueous extract of Limoniastrum guyonianum gall.

ETHNOPHARMACOLOGICAL RELEVANCE: Many studies have been performed to assess the potential utility of natural products as immunomodulatory agents to enhance host responses to disease/infection/etc. or to ameliorate immune based pathologies (i.e., inflammation, autoimmune associated diseases, etc.). In this particular study, the immunomodulatory potential of gall aqueous extract from Limoniastrum guyonianum Boiss. (Zita) was assessed in vitro. MATERIALS AND METHODS: The effect of G extract on splenocytes proliferation and NK activity were assessed by MTT test. The induction of NO production and the phagocytic activity of macrophages were evaluated in vitro. Activation of the cellular anti-oxidant activity in splenocytes was determined by measuring the fluorescence of the DCF product. RESULTS: The studies first demonstrated that the extract could enhance lysosomal enzyme activity and nitrite oxide production in murine peritoneal macrophages, suggesting a potential role in activation of these cells. In studies to assess potential effects on humoral immunity, the results indicated that the extract could significantly promote LPS-stimulated splenocyte proliferation implying a potential activation of B-cells and enhanced humoral immune responses in hosts given this natural product. In studies to assess any effects of extract on cellular immunity, the results showed that the extract significantly enhanced the killing activity of isolated NK cells but had negligible effects on mitogen-induced proliferation of splenic T-cells. Considerable effects were also observed on the cellular anti-oxidant activity. CONCLUSION: We conclude from these studies that aqueous extract from L. guyonianum gall exhibited an immunomodulator effect which could be ascribed, in part, to its cytoprotective effect via its anti-oxidant capacity. Furthermore, these results suggest that L. guyonianum gall extract contains potent components such as flavonoids which should be potentially used to modulate immune cell functions in physiological and pathological conditions.

Arginine-supplemented enteral nutrition in critically ill diabetic and obese rats: a dose-ranging study evaluating nutritional status and macrophage function.

OBJECTIVE: Critically ill diabetic and obese patients are at high risk of complications. Arginine availability is lowered in diabetes and in stress situations, yet arginine is necessary for immune response, mainly by its action through nitric oxide (NO). These facts argue for arginine-supplemented diets in critically ill patients. However, studies have raised concerns about possible adverse effects of such diets in intensive-care patients. We therefore analyzed the metabolic and immunologic effects of an arginine-enriched diet in stressed diabetic-obese rats. METHODS: Zucker Diabetic Fatty rats (fa/fa) were made endotoxemic by an intraperitoneal injection of lipopolysaccharide and then fed 4-d enteral nutrition enriched with arginine (ARG group) or a non-essential amino acid mix (NEAA group). The two groups each were subdivided into three subgroups: the ARG subgroups received 0.5 g (ARG0.5), 2 g (ARG2), and 5 g (ARG5) of arginine per kilogram daily, and the NEAA groups were made isonitrogenous with the corresponding ARG subgroups (NEAA0.5, NEAA2, and NEAA5). Plasma and urinary biomarkers were measured. Cytokine and NO production levels and inducible NO synthase and arginase protein levels were determined from peritoneal macrophages. RESULTS: The survival rate was lower in the ARG5 and NEAA5 subgroups than in all the other subgroups. The nitrogen balance was higher in the ARG5 group than in the NEAA5 group. Plasma triacylglycerol levels were lower in the ARG2 group than in the NEAA2 group. Interleukin-6, tumor necrosis factor-α, and NO production in the macrophages decreased and arginase-1 was upregulated in the ARG-treated rats. CONCLUSIONS: In this model, mortality was increased by the nitrogen burden rather than by arginine per se. Arginine improved nitrogen balance and had an anti-inflammatory action on macrophages by regulating NO production, probably through arginase-1 expression.

4-(1H-Pyrazol-1-yl) benzenesulfonamide derivatives: identifying new active antileishmanial structures for use against a neglected disease.

Leishmaniasis is a neglected disease responsible for about 56,000 deaths every year. Despite its importance, there are no effective, safe and proper treatments for leishmaniasis due to strain resistance and/or drug side-effects. In this work we report the synthesis, molecular modeling, cytotoxicity and the antileishmanial profile of a series of 4-(1H-pyrazol-1-yl)benzenesulfonamides. Our experimental data showed an active profile for some compounds against Leishmania infantum and Leishmania amazonensis. The profile of two compounds against L. infantum was similar to that of pentamidine, but with lower cytotoxicity. Molecular modeling evaluation indicated that changes in electronic regions, orientation as well as lipophilicity of the derivatives were areas to improve the interaction with the parasitic target. Overall the compounds represent feasible prototypes for designing new molecules against L. infantum and L. amazonensis.

Characterization and immunostimulating effects on murine peritoneal macrophages of oligosaccharide isolated from Panax ginseng C.A. Meyer.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax ginseng C.A. Meyer has been the most precious and renowned Chinese herb used in Asian countries for the treatment of various medical disorders. AIM OF THE STUDY: The aim of this work was to investigate the activation effect on murine peritoneal macrophages of oligosaccharide from the roots of P. ginseng. MATERIALS AND METHODS: In this work, the water-extracted oligosaccharide of P. ginseng was (WGOS) isolated and purified from the roots of P. ginseng by hot water extraction, ultrafiltration and gel-permeation chromatography. The monosaccharide composition and degree of polymerization (DP) of WGOS were determined by a combination of acid hydrolysis, high performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis. Phagocytosis of macrophages was measured by uptake of the neutral red by macrophages, nitric oxide (NO) was determined by the Griess method, inducible NO synthase (iNOS) activity was determined by colorimetric method using a reagent kit, and tumor necrosis factor-α (TNF-α) was analyzed by enzyme linked immunosorbent assay (ELISA). The reactive species detection kit was used to measure the reactive oxygen species (ROS) level. RESULTS: WGOS was composed of glucose and the DP was ranging from 2 to 14. Immunological tests showed that treatment of WGOS significantly increased phagocytosis of macrophages, and promoted NO, TNF-α and ROS production. Furthermore, WGOS dose-dependently stimulated NO formation through the up-regulation of iNOS activity. CONCLUSIONS: Taken together, WGOS possessed high immunopotentiating activity and could be developed as a novel immnunostimulant.

Further analysis of the structure and immunological activity of an RG-I type pectin from Panax ginseng.

In this paper, we further analysed the structure of a type I rhamnogalacturonan (RG-I) pectin (WGPA-2-RG) fractionated from ginseng polysaccharides. Methylation and periodate oxidation analyses showed that WGPA-2-RG has a backbone consisting of alternating rhamnose (Rha) and galacturonic acid (GalA) residues and side chains consisting of type II arabinogalactan (AG-II). Partial acidic hydrolysis for 6h completely removed arabinose (Ara), partial galactose (Gal), but little GalA and Rha. During partial hydrolysis, the molecular weight of WGPA-2-RG decreased smoothly, suggesting that the Ara and cleavable Gal residues exist on the surface of the molecule, while GalA and Rha residues exist in the core of the molecule. The bioactivity assay showed that the arabinogalactan side chains of WGPA-2-RG are essential structures for stimulating NO secretion and lymphocyte proliferation. However, removal of the Ara and Gal residues through hydrolysis did not appreciably affect the ability of WGPA-2-RG to enhance macrophage phagocytosis.

Effect of the Gc-derived macrophage-activating factor precursor (preGcMAF) on phagocytic activation of mouse peritoneal macrophages.

BACKGROUND: The 1f1f subtype of the Gc protein (Gc(1f1f) protein) was converted into Gc-derived macrophage-activating factor (GcMAF) by enzymatic processing in the presence of ß-galactosidase of an activated B-cell and sialidase of a T-cell. We hypothesized that preGc(1f1f)MAF, the only Gc(1f1f) protein lacking galactose, can be converted to GcMAF in vivo because sialic acid is cleaved by residual sialidase. Hence, we investigated the effect of preGc(1f1f)MAF on the phagocytic activation of mouse peritoneal macrophages. RESULTS: We examined the sugar moiety of preGc(1f1f)MAF with a Western blot using peanut agglutinin (PNA) and Helix pomatia agglutinin (HPA) lectin. We also found that preGc(1f1f)MAF significantly enhanced phagocytic activity in mouse peritoneal macrophages but only in the presence of the mouse peritoneal fluid; the level of phagocytic activity was the same as that observed for GcMAF. CONCLUSION: PreGc(1f1f)MAF can be used as an effective macrophage activator in vivo.

Solanum lyratum extract affected immune response in normal and leukemia murine animal in vivo.

Solanum lyratum Thunberg (Solanaceae) has been used as a folk medicine for treating liver, lung and esophagus in the Chinese population. Our previous studies have shown that the crude extract of S. lyratum Thunberg (SLE) induced apoptosis in colo 205 human colon adenocarcinoma cells; however, there is no report to show SLE affect immune responses in vivo. In this study, the in vivo effects of SLE on leukemia WEHI-3 cells and immune responses such as phagocytosis and natural killer (NK) cell activity in normal and leukemia mice were investigated. The SLE treatment decreases surface markers of CD3 and Mac-3 in normal and leukemia mice but promoted the cell markers of CD19 and CD11b in normal mice and CD11b in leukemia mice indicating that the precursors of T cells was inhibited and B cells and macrophage were promoted. The SLE treatment promoted the activity of macrophage phagocytosis in the peripheral blood mononuclear cells (PBMC) and peritoneal cells from normal and leukemia mice. The results also showed that NK cells from the normal and leukemia mice after treatment with SLE can kill the YAC-1 target cells. Therefore, the SLE treatment increased macrophage and NK cell activities. These consistent results indicate SLE could be a potent immune responses agent.