Jian Du

Ethyl acetate extraction from a Chinese herbal formula, Jiedu Xiaozheng Yin, inhibits the proliferation of hepatocellular carcinoma cells via induction of G0/G1 phase arrest in vivo and in vitro

Jiedu Xiaozheng Yin (JXY), a polyherbal formula of traditional Chinese medicine (TCM), has been used to treat various kinds of cancer in China. However, the mechanism of its anticancer activity has yet to be elucidated. Air-dried herbs were extracted with reagents of different polarity. HepG2 cells were treated with different doses of ethyl acetate extract (EE-JXY) and chloroform extract (CE-JXY) for 24 h. Cell viability was detected by MTT assay. Colony formation ability was also evaluated. Cell cycle was evaluated by FACS. Tumor bearing BALB/c nude mice was treated with EE-JXY (0.06 g/kg) for 20 days. Tumor volume and weight were monitored. The percentage of PCNA-positive cells and the level of G1 phase proteins [cyclin-dependent kinase2 (CDK2), cyclin‑dependent kinase4 (CDK4), cyclin D and cyclin E and G2 phase proteins [cyclin-dependent kinase1 (CDK1), cyclin A and cyclin B] were detected by immunohistochemistry and western blotting. EE-JXY and CE-JXY dose-dependently inhibited the growth of HepG2 cells (P<0.01 for both). Furthermore, EE-JXY inhibited the formation of cell colonies and blocked the cell cycle to G1 phase in a dose-dependent manner (P<0.01 for all). EE-JXY showed an obviously antitumor effect in vivo (P<0.05). Further investigation showed that EE-JXY decreased the proliferation index of tumors (P<0.01) through increasing the expression of G1-related proteins (cyclin D and cyclin E, P<0.05 and P<0.01). These results suggested that JXY inhibits the growth of HepG2 cells at least via arresting the cell cycle at the G0/G1 phase.

Traditional Chinese medicine syndrome-related herbal prescriptions in treatment of malignant tumors.

OBJECTIVE To investigate the distribution characteristics of TCM syndromes and the related herbal prescriptions for malignant tumors (MT). METHODS A clinical database of the TCM syndromes and the herbal prescriptions in treatment of 136 MT patients were established. The data were then analyzed using cluster and frequency analysis. RESULTS According to the cluster analysis, the TCM syndromes in MT patients mainly included two patterns: deficiency of both Qi and Yin and internal accumulation of toxic heat. The commonly-prescribed herbs were Huangqi (Astraglus), Nüzhenzi (Fructus Ligustri Lucid), Lingzhi (Ganoderma Lucidum), Huaishan (Dioscorea Opposita), Xiakucao (Prunella Vulgaris), and Baihuasheshecao (Herba Hedyotidis). CONCLUSION Deficiency of Qi and Yin is the primary syndrome of MT, and internal accumulation of toxic heat is the secondary syndrome. The herbs for Qi supplementation and Yin nourishment are mainly used, with the assistance of herbs for heat-clearance and detoxification.

Fuzheng Yiliu Granule (扶正抑瘤颗粒) inhibits the growth of hepatocellular cancer by regulating immune function and inducing apoptosis in vivo and in vitro

ObjectiveTo study the inhibitory effect of Fuzheng Yiliu Granule (扶正抑瘤颗粒, FYG) on hepatocellular cancer (HCC) and investigate the mechanism mediating its bioactivity.MethodsH22 tumor-bearing ICR mice were treated with FYG [3.6 g/(kg·d)] for 5 days. Tumor volume and tumor weight, percentages of CD3+, CD4+, CD8+, and natural killer (NK) cells in peripheral blood, tumor apoptosis and serum levels of interleukin-2 (IL-2), and tumor necrosis factor-α (TNF-α) were evaluated. FYG-containing serum was prepared from SD rats treated for 7 days [high dose 3.6 g/(kg·d); middle dose 1.8 g/(kg·d); low dose 0.9 g/(kg·d)]. Cell cycle, cell viability, and apoptosis were evaluated after HepG2 cell line was cultured in FYG-containing serum for 48 h. The levels of IL-2 and TNF-α in FYG-containing serum were also determined.ResultsFYG produced a potent antitumor effect (P<0.01) and induced marked apoptosis of the tumor tissue (P<0.05). Mice treated with FYG had higher percentages of CD3+ and CD4+ (P<0.05), and more NK cells (P<0.01) in the peripheral blood than those in the animals treated with normal saline. Mice receiving FYG had the highest serum levels of IL-2 and TNF-α (P<0.01). High-dose FYG-containing serum significantly decreased HepG2 cell viability, inhibited cell proliferation (P<0.05), and induced apoptosis (P<0.01). In addition, the levels of IL-2 and TNF-α of high-dose-containing serum were higher than the blank serum (P<0.01).ConclusionFYG could inhibit HCC growth by regulating immune function and inducing apoptosis of tumor cells in vivo and in vitro.

Enhancement of Antitumor Activity of Low-Dose 5-Fluorouracil by Combination With Fuzheng-Yiliu Granules in Hepatoma 22 Tumor-Bearing Mice

Objective. The adverse effects of 5-fluorouracil (5-FU) are well recognized. Fuzheng-Yiliu granule (FYG) is capable of enhancing the immune function and suppressing tumor growth. In the present study, the authors evaluated if FYG could synergize with low-dose 5-FU in inhibiting tumor growth. Methods. Hepatoma 22 (H22) tumor-bearing mice were treated with FYG (18 g/kg, ig), 5-FU (10 mg/kg, ip), or 5-FU plus FYG for 5 days. The relative tumor proliferation rates, tumor weight and apoptosis of tumor tissue were measured. White blood cell (WBC) and lymphocyte (LY) were counted. Interleukin-2 (IL-2) and tumor necrosis factor (TNF-a) in the serum were measured. Results. FYG alone had antitumor effect. Combination of 5-FU and FYG produced a more potent antitumor effect and caused more marked apoptosis in tumor tissue (compared with vehicle, P < 0.01; compared with 5-FU or FYG, P < 0.05). Mice treated with 5-FU plus FYG had higher thymus index (P < 0.05) compared with the vehicle group. The numbers of both WBC and LY were decreased by 5-FU (compared with vehicle, P < 0.01), which was significantly reversed after FYG was administered (5-FU + FYG vs 5-FU, P < 0.01 and P < 0.05). Mice receiving FYG alone or FYG plus 5-FU had higher serum levels of TNF-a (P <0.01) compared with the vehicle. Conclusions. Traditional Chinese medical herbs capable of strengthening the body’s vital energy have great potential to be used as an adjuvant therapy for cancer patients who cannot tolerate the adverse effects of chemotherapy.

Antitumor and immunomodulatory effects of low‑dose 5‑FU on hepatoma 22 tumor‑bearing mice

Low-dose 5-fluorouracil (5-FU), a widely used chemotherapeutic, has been reported to have immunomodulatory effects. This study aimed to evaluate the optimal dose of 5-FU that produces antitumor and immunomodulatory effects. In a hepatoma 22 tumor-bearing mouse model, 0, 10, 20 and 40 mg/kg 5-FU (i.p.) was administered for 10 days. Tumor weight and volume were measured, thymus index (TI) and spleen index (SI) were calculated, and the number of white blood cells (WBCs) and lymphocytes (LYs) were counted following treatment. The percentages of CD3+, CD4+, CD8+ and natural killer (NK) cells were measured by flow cytometry. In addition, the body weights of the mice were measured and the average diet consumption was calculated. Administration of 5-FU produced a potent antitumor effect in a dose-dependent manner (P<0.01). At 20 and 40 mg/kg, a significant reduction of body weight and food consumption was observed. TI and SI decreased in the 20- and 40-mg/kg groups (P<0.01) for 10 days. The number of WBCs significantly decreased in each group (P<0.01); however, the number of LYs only decreased in the 40-mg/kg group (P<0.01). Percentages of CD3+ and CD4+ cells were increased in the 10- and 20-mg/kg groups (P<0.01). Thus, 5-FU at 10 mg/kg inhibits tumor growth while maintaining the immune function of the mice. 5-FU may exert its antitumor effect at a low dose with low toxicity and stimulate the host immune system. Future clinical trials taking into account the immunostimulatory capacity of chemotherapeutic agents are desirable for certain patients.

Application of serum pharmacology in evaluating the antitumor effect of Fuzheng Yiliu Decoction (扶正抑瘤方) from Chinese medicine

ObjectiveTo investigate the feasibility of serum pharmacology in evaluating the antitumor effect of Chinese medicine (CM) of Fuzheng Guben (扶正固本, supporting the healthy energy and strengthening the body’s resistance to pathogens), the effects of Fuzheng Yiliu Decoction (扶正抑瘤方, FYD), a typical prescription of Fuzheng Guben, on proliferation and apoptosis of hepatoma cells in vitro were observed by two methods with serum pharmacology and traditional pharmacology, respectively.MethodsHepG2 cells were treated with FYD-containing serum or crude FYD extract in vitro. The proliferation rate was determined by methyl thiazolyl tetrazolium (MTT) assay. Cell cycle and apoptosis rate was performed by flow cytometry. And the levels of interleukin-2 (IL-2) and tumor necrosis factor α (TNF-α) in FYD-containing serum were detected by radioimmunoassay.Results FYD-containing serum remarkably inhibited proliferation and induced apoptosis of hepatoma cells at least by promoting the production of IL-2 and TNF-α in vivo . On the contrary, crude FYD extract promoted the proliferation and did not induce cell apoptosisConclusionThe results by serum pharmacology were accordant with those of our previous animal and clinical trials which indicates that serum pharmacology is a reasonable and feasible method for the evaluation of the antitumor effect of herbs of Fuzheng Guben.

Applied research on serum protein fingerprints for prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer.

OBJECTIVE This study screened serum tumor biomarkers by surface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) to establish a subset which could be used for the prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer; and as diagnostic model of Chinese medicine. METHODS Serum samples from 63 lung cancer patients with Qi deficiency syndrome and phlegm and blood stasis, and 28 lung cancer patients with non-Qi deficiency syndrome and phlegm and blood stasis were analyzed using SELDI-TOF-MS with a PBS II-C protein chip reader. Protein profiles were generated using immobilized metal affinity capture (IMAC3) protein chips. Differentially-expressed proteins were screened. Protein peak clustering and classification analyses were performed using Biomarker Wizard and Biomarker Pattern software packages, respectively. RESULTS A total of 268 effective protein peaks were detected in the 1,000-10,000 Da molecular range for the 15 serum proteins screened (P<0.05). The decision tree model was M 2284.97, with a sensitivity of 96.2% and a specificity of 66.7%. CONCLUSION SELDI-TOF-MS techniques, combined with a decision tree model, can help identify serum proteomic biomarkers related to Qi deficiency syndrome and phlegm and blood stasis in lung cancer patients; and the predictive model can be used to discriminate between Chinese medicine diagnostic models of disease.

Ethyl acetate extract from Jiedu Xiaozheng Yin inhibits the proliferation of human hepatocellular carcinoma cells by suppressing polycomb gene product Bmi1 and Wnt/β-catenin signaling

Jiedu Xiaozheng Yin (JXY) is a Chinese herbal decoction used to treat hepatocellular carcinoma (HCC). Previous studies have demonstrated that JXY can inhibit HCC cell proliferation via induction of G0/G1 phase arrest. In this study, we investigated whether the inhibitory effect of JXY on HCC cells is associated with the inhibition of the Wnt/β‑catenin pathway and the polycomb gene product Bmi1. Ethyl acetate extract from JXY (EE-JXY) was prepared. Methyl thiazolyl tetrazolium (MTT) and colony formation assays were used to measure cell proliferation. Immunofluorescence was used to analyze the expression and location of β-catenin and Bmi1. Immunohistochemistry was used to examine the expression of proliferating cell nuclear antigen (PCNA), c-myc and cyclin D1. β-catenin, Bmi1, c-myc, cyclin D1 and p16INK4A mRNA levels were detected by RT-PCR. The results demonstrated that EE-JXY inhibited the expression of PCNA, c-myc, cyclin D1 and Bmi1, and upregulated the expression of p16INK4A. We also found that EE-JXY could facilitate β-catenin translocation from the cytoplasm and nuclei to the cytomembrane. Finally, suppression of cell proliferation and expression of Bmi1 and Wnt/β-catenin by EE-JXY was confirmed in a mouse xenograft model of HCC. Thus, EE-JXY can inhibit the proliferation of HCC partially via suppression of the Bmi1 and Wnt/β-catenin signaling pathways.

Effective components of Chinese herbal compound decoction and Maillard reaction.

This paper intends to explore the color changes considered to be Maillard reaction during the process of Chinese herbal medicine. The Maillard reaction products (MRPs) are often in substantial proportions of Chinese herbal compound decoctions but their effects are often neglected. By considering the effects of MRPs in studies of effective components on Chinese herbal compounds, a new perspective is established in future researches of Chinese herbal compound decoctions.

Active hexose correlated compound potentiates the antitumor effects of low-dose 5-fluorouracil through modulation of immune function in hepatoma 22 tumor-bearing mice

BACKGROUND/OBJECTIVES A variety of immunomodulators can improve the efficacy of low-dose chemotherapeutics. Active hexose correlated compound (AHCC), a mushroom mycelia extract, has been shown to be a strong immunomodulator. Whether AHCC could enhance the antitumor effect of low-dose 5-fluorouracil (5-FU) via regulation of host immunity is unknown. MATERIALS/METHODS In the current study Hepatoma 22 (H22) tumor-bearing mice were treated with PBS, 5-FU (10 mg·kg-1·d-1, i.p), or AHCC (360 mg·kg-1·d-1, i.g) plus 5-FU, respectively, for 5 d. CD3+, CD4+, CD8+, and NK in peripheral blood were detected by flow cytometry. ALT, AST, BUN, and Cr levels were measured by biochemical assay. IL-2 and TNFα in serum were measured using the RIA kit and apoptosis of tumor was detected by TUNEL staining. Bax, Bcl-2, and TS protein levels were measured by immunohistochemical staining and mRNA level was evaluated by RT-PCR. RESULTS Diet consumption and body weight showed that AHCC had no apparent toxicity. AHCC could reverse liver injury and myelosuppression induced by 5-FU (P < 0.05). Compared to mice treated with 5-FU, mice treated with AHCC plus 5-FU had higher thymus index, percentages of CD3+, CD4+, and NK cells (P < 0.01), and ratio of CD4+/CD8+ (P < 0.01) in peripheral blood. Radioimmunoassay showed that mice treated with AHCC plus 5-FU had the highest serum levels of IL-2 and TNFα compared with the vehicle group and 5-FU group. More importantly, the combination of AHCC and 5-FU produced a more potent antitumor effect (P < 0.05) and caused more severe apoptosis in tumor tissue (P < 0.05) compared with the 5-FU group. In addition, the combination of AHCC and 5-FU further up-regulated the expression of Bcl-2 associated X protein (Bax) (P < 0.01), while it down-regulated the expression of B cell lymphoma 2 (Bcl-2) (P < 0.01). CONCLUSIONS These results support the claim that AHCC might be beneficial for cancer patients receiving chemotherapy.