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ObjectiveTo investigate the effect of Ganoderma lucidum polysaccharides （GLP） on the proliferation， migration， cycle， and apoptosis of hepatocellular carcinoma SKHEP1 and Huh7 cells and to explore the underlying mechanism. MethodSK-HEP-1 and Huh-7 cells were classified into the blank group and low-， medium-， and high-dose GLP groups （3.5， 7， 14 g·L-1）. The proliferation of the cells was examined by cell counting kit-8 （CCK8） assay， and the migration by scratch assay. Cell cycle was measured by flow cytometry and apoptosis was detected based on Hoechst33258 staining. In addition， the expression of phosphatidylinositol 3-kinase （PI3K）， protein kinase B （Akt）， phosphorylated PI3K （pPI3K）， and phosphorylated Akt （pAkt） in the cells was determined by Western blot. ResultCompared with the blank group， the three doses of GLP reduced the proliferation and migration of SKHEP1 and Huh7 cells （P<0.05）， increased the percentage of cells in G1 phase （P<0.05）， and decreased percentage of cells in S and G2 phase （P<0.05）. In addition， the three doses can induce apoptosis of both SK-HEP-1 and Huh-7 cells， particularly the high dose. Moreover， the three doses of GLP lowered the levels of pPI3K and pAkt （P<0.05）. ConclusionGLP significantly inhibited the malignant phenotype of SK-HEP-1 and Huh-7 cells through the PI3K/Akt signaling pathway.
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With the population aging， the morbidity and mortality of cancer patients continue to rise. At present， the treatment methods for tumors include surgery， chemotherapy， radiotherapy， targeted therapy， and immunotherapy. However， most chemotherapeutic drugs can cause severe side effects and drug resistance. Therefore， as an alternative therapy， traditional Chinese medicine （TCM） treatment can effectively relieve the clinical symptoms of tumor patients， improve the quality of life， inhibit or stabilize the development of tumors， and prolong the survival period of patients. Due to the good safety of Chinese medicine， its potential anti-cancer activity has attracted increasing attention. Ganoderma lucidum， a treasure of Chinese medicinal material， is a medicinal fungus with a history of more than 2 000 years in China. So far， many studies have proposed the anti-cancer properties of G. lucidum. G. lucidum has extensive pharmacological activities， such as anti-tumor， anti-atherosclerosis， and anti-aging. It can also regulate immunity， protect the liver and the heart， and reduce blood glucose and lipid. The chemical composition of G. lucidum is complex. At present， it is proved to contain polysaccharides， triterpenoids， alkaloids， nucleosides， amino acids， and various trace elements. The anti-tumor mechanisms of polysaccharides and triterpenoids in G. lucidum are mainly achieved by apoptosis induction， immune regulation， anti-angiogenesis， and induction of cell cycle arrest. Currently， it has been widely used in the adjuvant treatment of complex tumors such as lung cancer， liver cancer， cervical cancer， prostate cancer， breast cancer， and colon cancer. The present study reviewed the bioactivities and mechanisms of triterpenoids and polysaccharides in G. lucidum in recent years and highlighted the anti-tumor effects and mechanisms to provide references for the further development and utilization of G. lucidum.
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Polysaccharide is among the main active components of Ganoderma lucidum for tumor prevention and treatment. Howe-ver, it remains unclear whether it has synergy with tumor immunotherapy. This study evaluated the effect of G. lucidum polysaccharides(GLP) on the infiltration of T lymphocytes into tumor and the underlying mechanism, in order to provide a reference for its application in tumor immunotherapy. GLP were prepared by water extraction and alcohol precipitation combined with Sevag method and then given(intraperitoneal injection) to the mice bearing B16-F10 cells at 25, 50 and 100 mg kg~(-1), respectively, to evaluate the effect on tumor growth. The infiltration of CD3~+ and CD8~+ T cells and the expression of intercellular cell adhesion molecule-1(ICAM-1) in tumor were detected by immunohistochemistry. EA.hy926 cells were treated with 50, 100 and 200 μg·mL~(-1) GLP, and the expression of ICAM-1 was determined by Western blot. The adhesion of EA.hy926 cells treated with GLP was measured with fluorescence-labeled Jurkat cells. To analyze the mechanism based on NF-κB pathway, this study determined the protein levels of nuclear factor kappa-B(NF-κB) p65, alpha inhibitor of NF-κB(IκBα), p-NF-κB p65 and p-IκBα by Western blot. The results showed that GLP can significantly inhibit the tumor growth in mice bearing B16-F10 cells, promote the infiltration of CD3~+ and CD8~+ T cells in tumor, and increase the expression of ICAM-1 in tumor. Meanwhile, GLP could also enhance the expression of ICAM-1 in EA.hy926 cells, thus strengthen the adhesion to Jurkat cells, induce phosphorylation and protein degradation of IκBα, and raise the expression and phosphorylation level of NF-κB p65. These results suggested that GLP could promote the expression of ICAM-1 through NF-κB pathway and further enhance the infiltration of T lymphocytes into tumor, thereby inhibiting tumor growth. This study lays a foundation for the further application of GLP in tumor immunotherapy.
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Animals , Mice , Endothelial Cells/metabolism , Intercellular Adhesion Molecule-1/genetics , NF-kappa B/metabolism , Neoplasms , Polysaccharides , Reishi , Signal Transduction , T-Lymphocytes , Tumor Necrosis Factor-alphaABSTRACT
OBJECTIVE@#To investigate the effects of Ganoderma lucidum polysaccharides (GL-PS) on human fibroblasts and skin wound healing in Kunming male mice and to explore the putative molecular mechanism.@*METHODS@#Primary human skin fibroblasts were cultured. The viability of fibroblasts treated with 0, 10, 20, 40, 80, and 160 μg/mL of GL-PS, respectively were detected by 3-4,5-dimethyl-2-thiazolyl-2,5-diphenyl-2-Htetrazolium bromide (MTT). The migration ability of fibroblasts treated with 0, 10, 20, and 40 μg/mL of GL-PS were measured by transwell assay. The secretion of the C-terminal peptide of procollagen type I (CICP) and transforming growth factor-β1 (TGF-β1) in the cell supernatant was tested by enzyme-linked immunosorbent assay. The expression of β-catenin was detected by Western blot. Furthermore, the Kunming mouse model with full-layer skin resection trauma was established, and was treated with 10, 20, and 40 mg/mL of GL-PS, respectively as external use. The size of the wound was measured daily, complete healing time in each group was recorded and the percentage of wound contraction was calculated.@*RESULTS@#Compared with the control group, 10, 20, and 40 μg/mL of GL-PS significantly increased the viability of fibroblasts, promoted the migration ability of fibroblasts, and up-regulated the expressions of CICP and TGF-β1 in fibroblasts (Plt;0.05 or Plt;0.01). The expression of β-catenin in fibroblasts treated with 20 and 40 μg/mL of GL-PS was significantly higher than that of the control group (Plt;0.01). Furthermore, after external use of 10, 20, and 40 mg/mL of GL-PS, the rates of wound healing in mice were significantly higher and the wound healing time was significantly less than the control group (Plt;0.05 or Plt;0.01).@*CONCLUSION@#A certain concentration of GL-PS may promote wound healing via activation of the Wnt/β-catenin signaling pathway and up-regulation of TGF-β1, which might serve as a promising source of skin wound healing.
Subject(s)
Animals , Humans , Male , Mice , Cell Movement , Cell Survival , Cells, Cultured , Collagen Type I , Fibroblasts , Polysaccharides , Pharmacology , Reishi , Chemistry , Skin , Wounds and Injuries , Transforming Growth Factor beta1 , Physiology , Wound Healing , beta Catenin , PhysiologyABSTRACT
Objective Radiation can cause multiple damages to tissues and organs.This study aimed to explore the protec-tive effect of Ganoderma lucidum polysaccharides ( GLPs) against the 60 Co-γray radiation injury in mice and provide an experimental basis for the clinical use of GLPs. Methods One hundred mice were randomly divided into five groups of equal number: normal control, gavage control, radiation control, high-dose GLPs, and low-dose GLPs.Models of radiation injury were made in the mice by whole-body exposure to 60 Co-γrays.Three days before and after mod-eling, the animals in the high-dose and low-dose GLPs groups were given GLPs intragastrically at the dose of 100 and 50 mg/kg respec-tively, once daily for 14 days.Then the 30 day survival rate and sur-vival time of the model mice were recorded and the changes in the pe-ripheral blood index, spleen index, and serum superoxide dismutase( SOD) activity were observed. Results GLPs significantly increased the 30-day survival rate and the mean survival time of the mouse models (P<0.05), decreased the reduction of WBC count in the peripheral blood, and shortened the time of WBC restoration ( P<0.05 ) .Furthermore, GLPs obviously improved the spleen index and SOD activity of the Co-γray irradiated animals. Conclusion GLPs, with a significant anti-radiation effect, can effectively raise the survival rate of the mice exposed to a lethal dose of 60 Co-γrays, reduce radiation injury to WBC and platelets, and increase the activity of SOD in irradiated mice.
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Objective: To prepare Lingyi Formula multicomponent microemulsion and evaluate its anti-lung cancer activity. Methods: Lingyi Formula multicomponent microemulsion was prepared by aqueous titration method using polysaccharides solution of Ganoderma lucidum and Coix lachryma-jobi var. ma-yuen as aqueous phase and coix seed oil as oil phase, loading ganoderma triterpenes. The average particle size, Zeta potential, and stability were detected. The results of antitumor efficacy including tumor inhibitory rate, body weight change, immune organ index, and concentration of TNF-α and IL-6 were investigated. Besides, pathological section of tumor tissue and TUNEL labeling were conducted subsequently. Results: The prepared microemulsion displayed spherical surface with mean droplet size of (69.92 ± 8.43) nm, polydispersity (PDI) of 0.060 ± 0.008, and Zeta potential of (-11.30 ± 1.34) mV. Tumor inhibitory rate of microemulsion (57.25%) was significantly higher than that of suspension (45.89%), immune tissue index as well as the concentration of TNF-α and IL-6 were increased significantly. TUNEL labeling and pathological section of tumor tissue showed that the antitumor activity of microemulsion was significantly effective compared with that of suspensions. Conclusion: Lingyi Formula multicomponent microemulsion has a good anti-lung cancer activity.
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Aim To study the protective effects of Ganoderma lucidum polysaccharides peptide(GLPP)on ECV304 from oxidative injury.Methods Cultured ECV304 were injured by oxygen free radicals derived from tBOOH.Various concentrations of GLPP(12.5,25,50,100 mg?L-1)were added into culture medium.The survival rate of cells was measured by MTT assay.The morphological change of cells and injury of mitochondria were examined under the light and electron microscopes.The percentage of apoptosis of ECV304,labeled with AnnexinV/PI,was measured by flow cytometry.Results GLPP(12.5,25,50,100 mg?L-1)could reduce oxidative injury induced by tBOOH in ECV304 cells.The survival rate of cells treated with GLPP increased.The light microscopic examination showed that the injured cells decreased in GLPP-treated groups.Under the electron microscope it was found that GLPP(50 mg?L-1,incubated for 24 h)could protect the organelle such as mitochondria from oxidative injury and cells from apoptosis by tBOOH.The result of flow cytometry showed that the total percentage of apoptosis in control,GLPP and injury treated group was 2.24%?0.43%,24?6.4%(P
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Aim To study the effects of Ganoderma polysaccharide peptide (GLPP) on the production of nitric oxide (NO) in mice peritoneal macrophages induced by LPS and its mechanism. Methods The effects of GLPP on the NO releasing from mice peritoneal macrophages induced by LPS were detected by using Griess reagent and the expression of iNOS by GLPP in mice peritoneal macrophages was detected by immunohistochemical method.Result The production of NO was increased by GLPP.(25～200 mg?kg -1) ig for five days or by GLPP(3.125～200 mg?L -1) in vitro but the effects of LPS on the production of NO was not influenced significantly. The expression of iNOS was increased by GLPP(25～200 mg?kg -1) ig for five days or GLPP(3.125～200 mg?L -1) in vitro.Conclusion GPP in vivo or in vitro could increase the production of NO in mice peritoneal macrophages. It might take effects by enhancing the synthesis of iNOS.
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AIM: To investigate the free radical scavenging activity of Ganoderma lucidum polysaccharides peptide (GLPP) on peritoneal macrophages in mice. METHODS: Alloxan and tert butylhydroperoxide (tBOOH) were used as an oxidant to injury peritoneal macrophages in vivo in mice or in vitro, respectively. 2', 7' dichlorodihydrofluorescein diacetate (DCHF DA) was used as fluorescent probe. The fluorescence from cells was observed under the laser confocal microscope. Time series scan of conforcal microscope was used to observe the changes of fluorescence by GLPP in mice peritoneal macrophages over time. RESULTS: The results of confocal microscopy showed that GLPP (100 mg?kg -1 , ig for 5 d ) lowed fluorescence in the mice macrophages injured by alloxan (75 mg?kg -1 iv). GLPP (10 mg?L -1 ) also lowed fluorescence in the mice macrophages injured by tBOOH ( 7.76 ?10 -5 mol?L -1 ) in vitro as well. Time series scan showed that GLPP (10 mg?L -1 ) lowed fluorescence in the mice macrophages at rest state or during the respiratory burst induced by PMA (50 nmol?L -1 ). CONCLUSION: GLPP shows antioxidant effects and might have free radical scavenging effects on peritoneal macrophages in mice.
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AIM: To investigate the effect of (Ganoderma lucidum ) polysaccharides (Gl-PS) on the insulin-releasing function of pancreati c islets. METHODS: Pancreatic islets were isolated and incubated with 5.6 or 16.7 mmol?L -1 glucose and different concentratio ns of Gl-PS for 1 h. Then the insulin was examined. Verapamil, and verapamil co mbined EGTA were used to testify whether the insulin-releasing effect of Gl-PS was inhibited by these inhibitors. The effects of Gl-PS on protein expression of the glucose transporter 2(GLUT2) under 5.6 and 16.7 mmol?L -1 glucose were also investigated. RESULTS: Gl-PS stimulated th e insulin release when incubated with the glucose of 5.6 or 16.7 mmol ?L -1 . The insulin releasing effect of Gl-PS was partly inhibited by ve rapamil and completely inhibited by verapamil+EGTA. Gl-PS could promote the GLU T2 protein expression of pancreatic islets under glucose of 5.6 and 16.7 mmol?L -1 . CONCLUSION: Gl-PS possesses insulin-rele asing effect under the glucose of 5.6 and 16.7 mmol?L -1 due t o the promotion of the GLUT2 protein expression and the subsequent facilitation of Ca 2+ inflow into the pancreatic B cells.
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Aim To study the protective effects of Ganoderma lucidum polysaccharides peptide(GLPP)on the human umbilical vein endothelial cells(HUVECs)injured by reactive oxygen species(ROS),derived from tert-butylhydroperoxide(t-BOOH).Methods Vascular endothelial cells were obtained from infant umbilical cord for primary cultivation,identified by endothelial cell adhesion molecule-1(CD31)by fluorescence microscope.HUVECs were injured by ROS,derived from t-BOOH.The survival rate of cells was measured by Cell Counting Kit-8 assay.Hoechst 33258 nucleus staining was used to observe the early apoptosis injury by ROS.And the activation of Caspase-3 was detected by spectrophotometry.Electron microscopy was used to show the morphological changes in HUVECs.Results GLPP(6.125,12.5,25,50,100 mg?L-1)could inhibit the apoptosis of HUVECs by ROS.The survival rate of HUVECs was increased by GLPP and the percentage of cell apoptosis was decreased by it too.GLPP decreased the activation of Caspase-3 of HUVECs up-regulated by ROS.Electron microscope showed that the organelle such as mitochondria injury by ROS could be relieved by GLPP.Conclusion GLPP can prevent human umbilical vein endothelial cells from oxidative injury.