Preventive administration of juanbi capsules for knee osteoarthritis: effects on serum MMP-2 and MMP-9 levels and cartilage repair.

OBJECTIVE: To explore the mechanism of action of Juanbi Capsules, a Chinese medicine for invigorating the kidney and replenishing qi, in preventing osteoarthritis of the knee in rabbits. METHODS: Seventy-two 4-month-old, Japanese long-eared white rabbits were randomly divided into 6 groups: control (group A), model (group B), Chinese drug; high-dose (group C), Chinese drug; mid-dose (group D), Chinese drug; low-dose (group E), and drug control (group F). With the exception of the rabbits in group A, each rabbit was subjected to plaster cast fixation for 6 weeks to induce osteoarthritis. In addition, rabbits were administrated with an intragastric injection of the Chinese drug (groups C, D and E) or an aminoglucose hydrochloride capsule (group F) for 4 weeks. Blood was drawn from the central ear artery for serum MMP-2 and MMP-9 concentrations, and the knee joint cartilage was harvested for gross observation and light microscopy. RESULTS: There were significant differences in serum MMP-2 and MMP-9 concentrations between group B and groups C, D and E (P < 0.05), with no significant differences between groups D and F. Histological results showed various changes in tissue staining with treatment, with osteophyte and bone cyst formation, and superficial erosion in the articular surface of the cartilage; in some cases, the defect reached the mid-layer of the cartilage, and these changes were lower than those in the model group. CONCLUSION: Juanbi Capsules assist in preventing osteoarthritis in the rabbit, possibly by decreasing serum MMP-2 and MMP-9 levels.

Effects of preventive administration of juanbi capsules on TNF-alpha, IL-1 and IL-6 contents of joint fluid in the rabbit with knee osteoarthritis.

OBJECTIVE: To probe into the mechanism of the Chinese herbs with functions of reinforcing kidney and supplementing qi for preventing knee osteoarthritis of the rabbit. METHODS: Totally 72 healthy Japan long-ear white rabbits, aged 4 months, were randomly divided into 6 groups, blank group (A), model group (B), high dose Chinese herb group (C), middle dose Chinese herb group (D), small dose Chinese herb group (E), aminoglucose hydrochloride capsule control group (F), 12 rabbits in each group. All the rabbits in the groups, except the group A, were fixed with plaster cast for six weeks to establish rabbit knee osteoarthritis. At the same time of modeling, the different doses of Juanbi Capsules and aminoglucose hydrochloride capsule were administrated intragastrically in the group C, D, E, F, respectively, for 4 weeks, for preventive treatment. In the group B, the rabbit was administrated intragastrically with equal volume of normal saline to the medication groups, twice each day, in the morning and the evening, and in the group A, nothing was administrated. After modeling for 6 weeks, the joint fluid was taken and TNF-alpha, IL-1 and IL-6 contents were detected with ELISA method, and the articular cartilage was taken for macroscopic and microscopic examinations. RESULTS: In all the preventive treatment groups, the articular cartilage color changed to varying degrees with formation of osteophyte and bone cyst, superficial erosion on the chondral articular surface, and the cartilage defect reached to the mid layer in a part of specimens with cartilage exfoliation, but which in the extent were significantly lower than those in the model group. There were significant differences between the group A and B in TNF-alpha, IL-1 and IL-6 contents in the joint fluid (P < 0.05), indicating that the modeling is successful; and there were significant differences as group B compared with the group C,D, E, F, showing that TNF-alpha , IL-1 and IL-6 contents are decreased in all the medication groups; and significant differences between group C, D, E suggests that the increase of Chinese herb doses strengthened the effect of reducing TNF-alpha, IL-1 and IL-6 contents in joint fluid. CONCLUSION: The Juanbi Capsule prevents osteoarthritis possibly through decreasing serum TNF-alpha, IL-1 and IL-6 contents.

[Study on preparation of ampelopsin liposomes].

OBJECTIVE: To study the formulation and preparation of ampelopsin liposomes and evaluate their quality. METHOD: The liposomes were prepared by a film-ultrasonic dispersion technique. Served as quota with the entrapment ratio and appearance and diameter of the liposomes, the optimal formulation and preparation were selected by means of an uniform design test. The appearance of liposomes was observed by micrography. The diameter and electric charge of surface were determined by granularity mensuration instrument. The entrapment ratio and the leakage rate of ampelopsin liposome were determined by means of dialyze. The content of ampelopsin was determined by UV. RESULT: The result of electron micrography and the size distribution showed that the liposomes were similar to spherical small unilamellar vesicles. The mean diameter was (258.2 +/- 51.2) nm and the electric charge of surface is 19.0 mV. The entrapment ratio of ampelopsin liposomes was 62. 3% and the lecithoid oxidative rate was 0.83% (n = 3). CONCLUSION: The selected formulation and preparation of ampelopsin liposomes is efficient and practicable.

[Optimizing the extracting technique of ampelopsin from Ampelopsis cantoniensis Planch by a uniform design method].

OBJECTIVE: To optimize the preparation of ampelopsin from Ampelopsis Cantoniensis Planch. METHODS: The extraction and purification process was studied by the uniform design with the extract of ampelopsin content and purity as markers. The facters which influence the extraction and the purification of ampelopsin content were studied by uniform design. RESULTS: The optimum extraction and purification process: the concentration for alcohol was 90%, and refluxing quartic, 1.5 h each time; extraction by petroleum ether quintic, the mount of active carbon was 1 g/100 g of the medicine material, and recrystaling thrice. CONCLUSION: This extraction process has higher yield of ampelopsin and is available for production.

[The chemotaxis effect of ampelopsin on the immunocytes].

OBJECTIVE: To study the chemotaxis effect of ampelopsin with different concentration on monocytes and neutrophilic granulocytes. METHODS: Chemokinesis and chemotaxis tests were proceed in agarose gel comparing with chemokine IL-8 or MCP-1. RESULTS: At 25.6 microg/ml or 51.2 microg/ml, ampelopsin could strongly enhance the migration of neutrophilic granulocytes and monocytes. The chemotaxis effect induced by 25.6 microg/ml of ampelopsin had no significant differences with that induced by 150 ng/ml of IL-8 or 50 ng/ml of MCP-1 (P > 0.05). At a concentration of 12.8 microg/ml, the chemokime effect of ampelopsin was more potent than that of 150 ng/ml of IL-8 or 100 ng/ml of MCP-1 (P < 0.05). Ampelopsin exerted a synergistic action with IL-8 or MCP-1 on its chemotaxis effect to neutrophilic granulocytes and monocytes. CONCLUSION: Ampelopsin can strongly enhance the chemokinesis and chemotaxis effects of neutrophilic granulocytes and moncytes and exert a synergistic action with IL-8 or MCP-1 on its chemotaxis effect to neutrophilic granulocytes and monocytes.

[Inhibitory effects of ampelopsin on angiogenesis].

OBJECTIVE: To study the effect of ampelopsin on angiogenesis. METHODS: The anti-angiogenic effect was evaluated by MTT assay for proliferation of endothelial cells. The concentration of vascular endothelial growth factor (VEGF) and basic-fibroblast growth factor (bFGF) from human hepatocellular carcinoma Bel-7402 cells were detected by enzyme linked immunosorbant assay (ELISA). Immunohistochemical staining was conducted to detect the expression of VEGF and bFGF. The VEGF and bFGF in the cancer cells were examined by flow cytometry. The inhibitory effect of ampelopsin on the growth of human hepatocellular carcinoma Bel-7402 in nude mice was studied. RESULTS: Ampelopsin was shown to inhibit the proliferation of primary cultured bovine aortic endothelial cells in a concentration dependent manner in range of 6.4 - 51.2 microg/ml. The IC50 (50% inhibition concentration) value was 22.0 +/- 4.0 microg/ml. ELISA assay was shown that treatment with 12.8 microl/m1, 25.6 microl/ml and 38.4 microg/ml of ampelopsin resulted in an inhibition of VEGF production released by Bel-7402, and the inbibtitory rate was 14.2%, 40.0% and 49.6%, respectively. After exposure to 12.8 microg/ml of ampelopsin, a decrease in the expression and activity of VEGF and bFGF was observed by immunohistochemical staining. The concentration of VEGF and bFGF secretion by Bel-7402 cells were lower following ampelopsin treatment as shown by flow cytometry. Treatment with 25.6 microg/mL and 38.4 microg/ml of ampelopsin, the inbibitory rates were 32.2% and 57.4% for VEGF, and 54.9% and 62.6% for bFGF, respectively. The inhibitory rate of ampelopsin to the growth of the transplant tumor in nude mice were 24.3%, 41.4% and 45.75 respectively at the dose of 100 mg/kg, 150 mg/kg and 200 mg/kg. CONCLUSION: Ampelopsin is a potent inhibitor of VEGF and bFGF expression and production in human hepatocellular carcinoma Bel-7402 cell, and may be a promising angiogenesis inhibitor.

Ampelopsin, a small molecule inhibitor of HIV-1 infection targeting HIV entry.

OBJECTIVE: To investigate the anti-HIV effects of ampelopsin and its interaction with HIV-1 coreceptor CXCR4. METHODS: Through anti-virus experiments in vitro, the inhibitory effect of ampelopsin on HIV-1 infection was verified. Chemotaxis assay was performed to show the ability to induce PBMCs migration by ampelopsin, RANTES and SDF-1alpha. Fluorescence labelling monoclonal antibody was utilized to observe the interaction of ampelopsin and CXCR4. Mice immunosuppressant model was also established to detail the role ampelopsin played in regulating cellular immunological functions. RESULTS: Ampelopsin could protect sensitive cells against HIV-1 infection and dramatically reduce HIV-1 antigen P24 expression. HIV-1SF33 attaching to MT-4 cells was interfered by ampelopsin, and the EC50 was 0.175 mg/mL for cellular protection and 0.024 mg/mL for P24 inhibition. At co-cultivating phase, EC50 was 0.229 mg/mL and 0.197 mg/mL respectively. Furthermore, the EC50 was 0.179 mg/mL and 0.348 mg/mL in acute infection. Human PBMCs migration was induced after being challenged with ampelopsin or chemokines, and synergistic action was observed during co-treatment. Ampelopsin alone resulted in maximal chemotaxis at 1 mg/mL. HIV-1 co-receptor CXCR4 on the surface of PBMCs was decreased by internalization, which indicated the effect of ampelopsin on CXCR4. About 70% CXCR4 was reduced by ampelopsin at 1 mg/mL. Ampelopsin also augmented cellular immunological functions in immunosuppressive mice. CONCLUSION: Ampelopsin displays a strong inhibitive role during HIV-1 absorption, incubation and acute infection. These results are coincident with its immune enhancement.

[Effects of ampelopsin on invasion and metastasis of B16 mouse melanoma in vivo and in vitro].

OBJECTIVE: To investigate the effects of ampelopsin on B16 melanoma's invasion and metastasis in vivo and in vitro. METHOD: B16 mouse melanoma cells were injected into C57BL/6 mouse via tail lateral vein, which subsequently colonized into the animal lungs to form an experimental pulmonary metastasis of tumor cell. The ampelopsin was administered at 3 dosages by intraperitoneal injection daily for 18 days from the day before the cells injection. The B16 mouse melanoma cells were exposed to ampelopsin for 3 days. The effects of ampelopsin on invasion, migration and adhesion of B16 melanoma cells were evaluated with Transwell chambers or attachment with polycarbonate filters and reconstituted basement membrane (Matrigel). RESULT: The number of metastases in the animals that were given ampelopsin 150, 200, and 250 mg x kg(-1) x d(-1) was significantly reduced as compared to the vehicle control (P<0.05), and the inhibition rates were 30.97%, 40.58%, and 61.16%, respectively. The ability of the ampelopsin treated B16 cells to invade the reconstituted basement membrane was decreased significantly (P<0.01), and the inhibition rates were 36.06%, 59.58%, and 79.09% at 20 micromol x L(-1), 40 micromol x L(-1) and 80 micromol x L(-1) concentration, respectively. Ampelopsin could also inhibit B16 cells migration through PVPF in the Transwell chambers, and the inhibition rates were 51.59%, 56.51%, and 66.75% at 20 micromol x L(-1), 40 micromol x L(-1) and 80 micromol x L(-1), respectively (P<0.01). The ability of adhesion of the B16 cells by ampelopsin treated cells on fibronectin, laminin, or Matrigel was decreased significantly. CONCLUSION: Ampelopsin has anti-invasive and anti-metastatic effects on B16 melanoma.