Characterization of inulin-type fructans from two species of Radix Codonopsis and their oxidative defense activation and prebiotic activities.

BACKGROUND: Codonopsis pilosula and C. tangshen are both plants widely used in traditional Chinese medicine. Polysaccharides, which are their primary active components, are thought to be important in their extensive use. In this study, two neutral polysaccharide fractions of C. pilosula (CPPN) and C. tangshen (CTPN) were obtained by fractionation on a DEAE-Sepharose column and characterized. RESULTS: It was confirmed that the neutral polymers CPPN and CTPN were ß-(2,1)-linked inulin-type fructans with non-reducing terminal glucose, and degree of polymerization (DP) of 19.6 and 25.2, respectively. The antioxidant and prebiotic activities in vitro were assayed based on IPEC-J2 cell lines and five strains of Lactobacillus. Results indicated that the effects of CPPN and CTPN were increased antioxidant defense in intestinal epithelial cells through enhanced cell viability, improved expression of total antioxidant capacity, glutathione peroxidase, superoxide dismutase and catalase, and reduced levels of malondialdehyde and lactic dehydrogenase. The prebiotic activity of CPPN and CTPN was demonstrated by the promoting effect on Lactobacillus proliferation in vitro. The different biological activities obtained between the two fractions are probably due to the different DP and thus molecular weights of CPPN and CTPN. CONCLUSION: The inulin fractions from C. pilosula and C. tangshen were natural sources of potential intestinal antioxidants as well as prebiotics, which will be valuable in further studies and new applications of inulin-containing health products. © 2020 Society of Chemical Industry.

Protective Effects of Salidroside against Carbon Tetrachloride (CCl4)-Induced Liver Injury by Initiating Mitochondria to Resist Oxidative Stress in Mice.

The antioxidant effect of salidroside has been proven, but its role in liver injury is poorly understood. In this study, we aimed to evaluate the protective effects and mechanism of salidroside on liver injury induced by carbon tetrachloride (CCl4) in vivo. Mice were pretreated with salidroside (60 mg/kg, intraperitoneally injected, i.p.) once per day for 14 consecutive days and then administered with CCl4 (15.95 g/kg, i.p.) for 24 h to produce a liver injury model. Salidroside attenuated hepatic transaminase elevation in serum and ameliorated liver steatosis and necrosis, thereby suggesting its protective effect on the liver. Salidroside antagonized CCl4-induced toxicity by equilibrating antioxidation system, thereby inhibiting reactive oxygen species accumulation, and restoring mitochondrial structure and function. Salidroside exerts antioxidant and liver-protective effects by selectively inhibiting the activation of genes, including growth arrest and DNA -damage-inducible 45 α (Gadd45a), mitogen-activated protein kinase 7 (Mapk7), and related RAS viral oncogene homolog 2 (Rras2), which induce oxidative stress in the mitogen-activated protein kinase pathway. These results revealed that salidroside can protect the liver from CCl4-induced injury by resisting oxidative stress and protecting mitochondrial function.

A Polysaccharide Isolated from Codonopsis pilosula with Immunomodulation Effects Both In Vitro and In Vivo.

In this study, an acidic polysaccharide from Codonopsis pilosula Nannf. var. modesta (Nannf.) L. T. Shen (WCP-I) and its main fragment, WCP-Ia, obtained after pectinase digestion, were structurally elucidated and found to consist of a rhamnogalacturonan I (RG-I) region containing both arabinogalactan type I (AG-I) and type II (AG-II) as sidechains. They both expressed immunomodulating activity against Peyer's patch cells. Endo-1,4-ß-galactanase degradation gave a decrease of interleukine 6 (IL-6) production compared with native WCP-I and WCP-Ia, but exo-α-l-arabinofuranosidase digestion showed no changes in activity. This demonstrated that the stimulation activity partly disappeared with removal of ß-d-(1→4)-galactan chains, proving that the AG-I side chain plays an important role in immunoregulation activity. WCP-Ia had a better promotion effect than WCP-I in vivo, shown through an increased spleen index, higher concentrations of IL-6, transforming growth factor-ß (TGF-ß), and tumor necrosis factor-α (TNF-α) in serum, and a slight increment in the secretory immunoglobulin A (sIgA) and CD4+/CD8+ T lymphocyte ratio. These results suggest that ß-d-(1→4)-galactan-containing chains in WCP-I play an essential role in the expression of immunomodulating activity. Combining all the results in this and previous studies, the intestinal immune system might be the target site of WCP-Ia.

The Polysaccharides from Codonopsis pilosula Modulates the Immunity and Intestinal Microbiota of Cyclophosphamide-Treated Immunosuppressed Mice.

Based on previous studies about microflora regulation and immunity enhancement activities of polysaccharides from Codonopsis pilosula Nannf. var. modesta (Nannf.) L. T. Shen (CPP), there is little study on intestinal mucosal immunity, which is a possible medium for contacting microflora and immunity. In the present study, the BALB/c mice were divided into five groups (eight mice in each group), including a normal group (Con), a model control group (Model), and model groups that were administered CPP (50, 100, 200 mg/kg/d) orally each day for seven days after intraperitoneal injection of 60 mg/kg BW/d cyclophosphamide (CP) for three days. CPP recovered the spleen index and restored the levels of IFN-γ, IL-2, IL-10, as well as serum IgG. In addition, it elevated ileum secretory immunoglobulin A (sIgA), the number of Lactobacillus and acetic acid content in cecum. These results indicated that CPP plays an important role in the protection against immunosuppression, especially mucosa immune damage, and the inhibition of pathogenic bacteria colonization, which could be considered a potential natural source of immunoregulator.

Purification and Partial Structural Characterization of a Complement Fixating Polysaccharide from Rhizomes of Ligusticum chuanxiong.

Rhizome of Ligusticum chuanxiong is an effective medical plant, which has been extensively applied for centuries in migraine and cardiovascular diseases treatment in China. Polysaccharides from this plant have been shown to have interesting bioactivities, but previous studies have only been performed on the neutral polysaccharides. In this study, LCP-I-I, a pectic polysaccharide fraction, was obtained from the 100 °C water extracts of L. chuangxiong rhizomes and purified by diethylaminethyl (DEAE) sepharose anion exchange chromatography and gel filtration. Monosaccharide analysis and linkage determination in addition to Fourier transform infrared (FT-IR) spectrometer and Nuclear magnetic resonance (NMR) spectrum, indicated that LCP-I-I is a typical pectic polysaccharide, with homo-galacturonan and rhamnogalacturonan type I regions and arabinogalactan type I and type II (AG-I/AG-II) side chains. LCP-I-I exhibited potent complement fixation activity, ICH50 of 26.3 ± 2.2 µg/mL, and thus has potential as a natural immunomodulator.

Xiang-Qi-Tang and its active components exhibit anti-inflammatory and anticoagulant properties by inhibiting MAPK and NF-κB signaling pathways in LPS-treated rat cardiac microvascular endothelial cells.

Xiang-Qi-Tang (XQT) is a Chinese herbal formula containing Cyperus rotundus, Astragalus membranaceus and Andrographis paniculata. Alpha-Cyperone (CYP), astragaloside IV (AS-IV) and andrographolide (AND) are the three major active components in this formula. XQT may modulate the inflammatory or coagulant responses. We therefore assessed the effects of XQT on lipopolysaccharide (LPS)-induced inflammatory model of rat cardiac microvascular endothelial cells (RCMECs). XQT, CYP, AS-IV and AND inhibited the production of tumor necrosis factor alpha (TNF-α), intercellular cell adhesion molecule-1 (ICAM-1) and plasminogen activator inhibitor-1 (PAI-1), and up-regulated the mRNA expression of Kruppel-like factor 2 (KLF2). XQT and CYP inhibited the secretion of tissue factor (TF). To further explore the mechanism, we found that XQT, or its active components CYP, AS-IV and AND significantly inhibited extracellular signal-regulated kinase (ERK), c-jun NH2-terminal kinase (JNK) and p38 phosphorylation protein expression as well as decreased the phosphorylation levels of nuclear factor κB (NF-κB) p65 proteins in LPS-stimulated RCMECs. These results suggested that XQT and its active components inhibited the expression of inflammatory and coagulant mediators via mitogen-activated protein kinase (MAPKs) and NF-κB signaling pathways. These findings may contribute to future research on the action mechanisms of this formula, as well as therapy for inflammation- or coagulation-related diseases.

Effects of Jin-Ying-Tang on Staphylococcus aureus-induced mastitis in rabbit.

The present study was performed to investigate the effects of Jin-Ying-Tang (JYT), a Chinese herbal formula containing Lonicera japonica, Herba taraxaci, Fructus trichosanthis, Fructus forsythia, Radix et rhizoma rhei, Astragalus membranaceus, Angelica sinensis, on rabbit mastitis induced by Staphylococcus aureus. Suckling rabbits were challenged with 1.5 × 10(7) colony forming unit (CFU) of S. aureus at the base of the third pair teats, and they were treated and pretreated with JYT to detect the formula effects. The results showed that JYT could reduce the occurrence of Staphylococcal mastitis in rabbit model. To further investigate the action mechanism of JYT, we examined the leukocyte counts and inflammatory mediator levels such as TNF-α and IL-6 in blood and infected tissue. From histological study and blood analysis, we found that JYT could suppress leukocyte infiltration in infected mammary gland tissue and significantly inhibit the total leukocyte counts and lymphocytes (LYM), monocytes (MON) and granulocytes (GRA) fractions of leukocyte counts in blood. Enzyme-linked immunosorbent assay (ELISA) results showed JYT significantly decreased the TNF-α and IL-6 concentrations in serum and mammary gland. The analysis of these data suggested that JYT effectively inhibited inflammatory responses to reduce the occurrence of mastitis in rabbit model.

Xiang-qi-tang increases avian pathogenic Escherichia coli-induced survival rate and regulates serum levels of tumor necrosis factor alpha, interleukin-1 and soluble endothelial protein C receptor in chicken.

Xiang-qi-tang (XQT) is a Chinese herbal formula containing rhizoma Cyperi, Andrographis paniculata and Astragalus membranaceus. The present study investigated the effects of XQT on the mortality and inflammatory mediators in a chicken model challenged with avian pathogenic Escherichia coli (APEC). To detect the effect of XQT, the chickens were pretreated with the formula 12 h before being challenged with 10(8) colony forming unit (CFU) of APEC. The results showed that 0.6 g/kg XQT significantly elevated the survival rate of infected chickens. To further investigate the mechanism of decreasing mortality of XQT, we examined plasma inflammatory mediator levels. The levels of tumor necrosis factor alpha (TNF-α), interleukin-1 (IL-1) and soluble endothelial protein C receptor (sEPCR) were significantly increased in chickens challenged with APEC alone, whereas chickens pretreated with 0.6 g/kg XQT showed marked decrease of these inflammatory mediator levels during the death peak. Taken together, this study demonstrates that XQT has protective effects in APEC-treated chickens. The action mechanisms of XQT involve anti-inflammation and antithrombotic activity. These findings may contribute to future research on the action mechanisms of this formula, as well as prevention of or therapy for avian colibacillosis.

Purification, chemical characterization and antioxidant activities of polysaccharides isolated from Mycena dendrobii.

Two polysaccharides, MDP-1 and MDP-2, were obtained from the fermentation liquid of M. dendrobii by anion exchange chromatography and gel filtration. Their chemical structures were measured by FT-IR, GC, 1H and 13C NMR spectra, indicating the mainly compositions of mannose, xylose, and galactose for MDP-1; galacturonic acid, galactose and rhamnose for MDP-2. Furthermore, the antioxidant activities of MDPs were investigated, showing different antioxidant activities, in which MDP-2 performed noticeable, with excellent superoxide radical activity better than BHT, high DPPH radical activity (IC50 at 227 µg/mL) comparable with BHT, moderate reducing power activity and hydroxyl radical scavenging activity. The results indicated that the fermentation liquid of M. dendrobii could be used as a potential natural source of antioxidant.

Pyruvate Kinase M2 Activates mTORC1 by Phosphorylating AKT1S1.

In cancer cells, the mammalian target of rapamycin complex 1 (mTORC1) that requires hormonal and nutrient signals for its activation, is constitutively activated. We found that overexpression of pyruvate kinase M2 (PKM2) activates mTORC1 signaling through phosphorylating mTORC1 inhibitor AKT1 substrate 1 (AKT1S1). An unbiased quantitative phosphoproteomic survey identified 974 PKM2 substrates, including serine202 and serine203 (S202/203) of AKT1S1, in the proteome of renal cell carcinoma (RCC). Phosphorylation of S202/203 of AKT1S1 by PKM2 released AKT1S1 from raptor and facilitated its binding to 14-3-3, resulted in hormonal- and nutrient-signals independent activation of mTORC1 signaling and led accelerated oncogenic growth and autophagy inhibition in cancer cells. Decreasing S202/203 phosphorylation by TEPP-46 treatment reversed these effects. In RCCs and breast cancers, PKM2 overexpression was correlated with elevated S202/203 phosphorylation, activated mTORC1 and inhibited autophagy. Our results provided the first phosphorylome of PKM2 and revealed a constitutive mTORC1 activating mechanism in cancer cells.

Therapeutic effect of Chinese patent medicine "Wuhuanghu" on porcine infectious pleuropneumonia and its acute and subchronic toxicity as well as evaluation of safety pharmacology.

Chinese patent medicines play an important role in veterinary clinical use. The aim of this study is to research the anti-infection effect of Chinese patent medicine "Wuhuanghu" for the treatment of porcine infectious pleuropneumonia and to evaluate the safety of "Wuhuanghu" in order to provide a comprehensive understanding of its toxicity. The anti-infection results showed that the treatment with "Wuhuanghu" could significantly inhibit pneumonia and decrement of the pneumonia in high, medium and low doses of "Wuhuanghu" groups were 70.97%, 61.29% and 58.06% respectively. The acute toxicity test showed that rats in the highest group (5000mg/kg) had no death and no abnormal response, suggesting the LD50 of "Wuhuanghu" was more than 5000mg/kg. The subchronic toxicity study showed that hematology indexes in all groups had no obvious differences; blood biochemical index, only albumin and total cholesterol in middle and low doses of "Wuhuanghu" groups were significantly decreased when compared with control group. The clinical pathology showed that the target organ of "Wuhuanghu" was liver. The safety pharmacology study indicated that "Wuhuanghu" had no side effects on rats. In conclusion, "Wuhuanghu" has therapeutic and protective effects to porcine infectious pleuropneumonia in a dose-dependent manner and "Wuhuanghu" is a safe veterinary medicine.

Studies on the acaricidal mechanism of the active components from neem (Azadirachta indica) oil against Sarcoptes scabiei var. cuniculi.

Octadecanoic acid-3,4-tetrahydrofuran diester, isolated from neem (Azadirachta indica) oil, exhibited potent acaricidal activity against Sarcoptes scabiei var. cuniculi. In this paper, the acaricidal mechanism of octadecanoic acid-3,4-tetrahydrofuran diester against Sarcoptes scabiei var. cuniculi was evaluated based on pathologic histology and enzyme activities. The results showed that after compound treatment for 24h at a concentration of 20mg/mL, the lesions of mites were prominent under transmission electron microscopy. The lesions consisted of the lysis of dermis cell membranes and cell nuclear membranes, mitochondrial morphological abnormalities, the drop of spinal disorders, and mitochondrial vacuolization. The activity of superoxide dismutase (SOD), peroxidase (POD), glutathione-s-transferases (GSTs), and Ca(2+)-ATPase of mites significantly changed after treatment with octadecanoic acid-3,4-tetrahydrofuran diester compared with the control group. The activities of SOD, POD, and Ca(2+)-ATPase were significantly suppressed, whereas that of GSTs was activated. These results indicated that the mechanism of the acaricidal activity of octadecanoic acid-3,4-tetrahydrofuran diester was mainly achieved through interference with the energy metabolism of mites, thus resulting in insect death.

Acute and subacute toxicity study of 1,8-cineole in mice.

The effects of acute and subacute toxicity of 1,8-cineole in Kunming mice were studied. After acute oral administration, the LD50 value (95% CL) was 3849 mg/kg (3488.8~4247.1 mg/kg). In the subacute toxicity study, there were no significant differences in body weight and relative organ weight between the control group and 1,8-cineole treatment groups. The histopathological examinations showed that granular degeneration and vacuolar degeneration appeared in liver and kidney tissue after administration of high dose of 1,8-cineole. Under electron microscopy, a series of ultrastructural changes were observed: The electron microscopy assays indicated that the influence of 1,8-cineole on the target organ at the subcellular level were mainly on the mitochondria, endoplasmic reticulum and other membrane type structure of liver and kidney.

Sulfated derivatives of 20(S)-ginsenoside Rh2 and their inhibitory effects on LPS-induced inflammatory cytokines and mediators.

Ginsenoside Rh2 is one of the most important ginsenosides in ginseng with antitumor, antidiabetic, antiallergic, and anti-inflammatory effects. However, the extremely poor oral bioavailability induced by its low water solubility greatly limits the potency of Rh2 in clinical use. Therefore, in this study we sulfated 20(S)-ginsenoside Rh2 with chlorosulfonic acid and pyridine method, and got two new sulfated derivatives, Rh2-B1 and Rh2-B2, with higher water solubility. Their chemical structures were characterized by spectroscopic methods (IR, MS and NMR). Additionally, Rh2-B1 and Rh2-B2 had the greater anti-inflammatory effects than Rh2 through inhibiting inflammatory cytokines and mediators in LPS-induced mouse RAW264.7 macrophages cells. These results suggested that the sulfated modification of Rh2 improved its water solubility and the sulfated derivatives could be more potential candidates for developing as anti-inflammatory agents.

Toxicological evaluation of neem (Azadirachta indica) oil: acute and subacute toxicity.

Neem (Azadirachta indica), popularly known as traditional medicine is a native plant in India. Neem oil is a vegetable oil derived from seeds or fruits of the neem tree through pressing or solvent extraction, and largely used in popular medicine to have antifungal, antibacterial, antimalarial, antiparasitic, anti-inflammatory, as well as immunemodulatory properties in different animal species. In the present study, acute and 28-day subacute toxicity tests were carried out. In the acute toxicity test, the LD50 values of neem oil were found to be 31.95g/kg. The subacute treatment with neem oil failed to change body weight gain, food and water consumption. Serum biochemistry analysis showed no significant differences in any of the parameters examined under the dose of 1600mg/kg/day. Histopathological exams showed that the target organs of neem oil were testicle, liver and kidneys up to the dose of 1600mg/kg/day.