[Ruyiping Inhibited Pulmonary Metastasis of Breast Cancer by Regulating the Formation of Pre-metastasis Microenvironment].

Objective To observe the effect of Ruyiping (RYP, a recipe for fighting against re- currence and metastasis of breast cancer) on pre-metastatic microenvironment, and to study its possi- ble mechanism. Methods The experiment was divided into two parts. The 1st part lies in setting the pre- cancerous transfer, and the 2nd part lies in the effect of RYP on pre-metastatic microenvironment. There were 24 BALB/c mice in the 1st part. Logarithmic phase 4T1 cells were dispensed into cell suspension. Blood cells were counted by blood cell counter. Then they were injected into the 4th mammary fat pad of the 24 BALB/c mice under aseptic condition (1 x 106 cells/mL, 0.1 mL for each mouse). There were 60 BALB/c mice in the 2nd part. They were divided into the blank group, the model group, low, middle, high dose RYP groups by random digit table, 12 in each group. The modeling method was the same as men- tioned above. Medication was started from the 2nd day of inoculation. Mice in low, middle, high dose RYP groups were administered with 5. 13, 10. 26, 20. 52 g/kg RYP crude drugs per day by gastrogavage, once per day for 14 successive days. Equal volume of normal saline was administered by gastrogavage to mice in the blank group and the model group. Six mice were sacrificed at day 10, 14, 18, and 22, respectively in the 1 st part of the experiment. The pulmonary metastasis was observed. The histology and mi- cromorphology of lung tissues were observed under light microscope and electron microscope/transmission electron microscopy (TEM) in the 2nd part of the experiment. The relative pulmonary vascular per- meability was determined by Evans blue. The effect of RYP on the formation of pre-metastatic microenvironment was observed. The levels of angiogenin2 (Angpt2), vascular endothelial growth factor (VEGF) , IL6 and IL1 ß were detected by Western blot and Real time PCR. Results The period from day 0 to day 14 was considered to be the pre-metastatic phase. Compared with the model group, significant inhibition on the tumor weight and tumor volume were shown in middle and high dose RYP groups (P <0. 05,P <0. 01). RYP dose-dependently inhibited the tumor weight and tumor volume (P <0. 05,P <0. 01). Infiltration of lymphocytes occurred in the model group and the low dose RYP group. But there was no statistical difference in the morphology of lung tissue in light microscopic results between middle/high dose RYP groups and the blank group. The pulmonary blood vessel net was consisted of continuously densely capillaries. The structure of pulmonary capillaries was normal in the blank group. The blood vessel walls were not regular and even in the model group, with obviously distended capillaries. After treated by RYP, the injury was improved, with normal basic morphology of blood vessels. Compared with the blank group, the exudate in Evans blue was obviously increased, protein and mRNA expressions of Angpt2, VEGF, IL6, and IL1ß were increased in the model group (P <0. 05,P <0. 01). Compared with the model group, the exu- date in Evans blue was obviously decreased in each YRP group. The reduction of the exudate was dose- dependently with the dose of YRP (P <0. 01). Protein and mRNA expressions of VEGF in the middle dose RYP group, protein and mRNA expressions of Angpt2, VEGF, IL6, and ILI1ß were decreased in middle and high dose RYP groups (P <0. 05,P <0. 01). Protein expressions of IL6 were decreased in the middle dose RYP group (P <0. 01). Conclusions RYP had favorable regulation in the tumor growth and the formation of pre-metastatic microenvironment. It could protect the integrity of vascular system, inhibit the formation of pre-metastatic microenvironment possibly through inhibiting the expressions of Angpt2, VEGF, IL6, and IL11ß, and finally inhibiting the occurrence of pulmonary metastasis of breast cancer.

[Effects of platycodin D in combination with different active ingredients of Chinese herbs on proliferation and invasion of 4T1 and MDA-MB-231 breast cancer cell lines].

OBJECTIVE: To investigate the effects of platycodin D in combination with different active ingredients of Chinese herbs under different therapeutic principles on proliferation and invasion of 4T1 and MDA-MB-231 breast cancer cell lines. METHODS: The effective doses of platycodin D, Ophiopogon total saponins, curcumenol and osthole in inhibiting proliferation of breast cancer cell lines 4T1 and MDA-MB-231 were detected by methyl thiazolyl tetrazolium (MTT) assay, respectively. Optimized combinations of platycodin D with Ophiopogon total saponins, curcumenol, or osthole were determined by uniform design method. Effects of the optimized combinations of platycodin D with the three ingredients on proliferation and invasion of 4T1 and MDA-MB-231 cells were verified and evaluated by MTT assay and Transwell chamber test, respectively. RESULTS: Verifying study showed that the inhibitory effects of platycodin D in combination with curcumenol or osthole on proliferation of 4T1 and MDA-MB-231 cells were better than those of platycodin D in combination with Ophiopogon total saponins and each ingredient used alone (P<0.05 or P<0.01). The inhibitory effect of platycodin D in combination with Ophiopogon total saponins or osthole on invasion of 4T1 cells was significantly better than those of platycodin D in combination with curcumenol and each ingredient used alone (P<0.05 or P<0.01). Moreover, the inhibitory effect of platycodin D in combination with curcumenol or osthole on invasion of MDA-MB-231 cells was significantly better than that of platycodin D in combination with Ophiopogon total saponins (P<0.01). CONCLUSION: The optimized combinations of platycodin D with three different active ingredients of Chinese herbs under different therapeutic principles can significantly inhibit the proliferation and decrease the invasion of 4T1 and MDA-MB-231 cells. Different platycodin D combinations have different potency in suppressing breast cancer cell proliferation and invasion.

[Inhibitory effects of osthole, psoralen and aconitine on invasive activities of breast cancer MDA-MB-231BO cell line and the mechanisms].

OBJECTIVE: To explore the inhibitory effects and to investigate the mechanisms of combined treatment of osthole, psoralen with aconitine on human breast cancer cell line MDA-MB-231BO. METHODS: The best inhibitory concentration of osthole, psoralen combined with aconitine on MDA-MB-231BO cells was obtained by stepwise regression analysis after adopting a uniform experiment design. The invasive activities were observed by transwell assays, and expressions of transforming growth factor-ß1 (TGF-ß1), Smad2, Smad3, Smad4, Smad7, nuclear factor-κB (NF-κB) and receptor activator of NF-κB ligand (RANK) mRNAs were analyzed by real-time quantitative polymerase chain reaction. RESULTS: The optimal combination concentrations of osthol, psoralen and aconitine were 6.44, 8.89 and 9.44 µg/mL, respectively. Cell invasion was significantly inhibited after 24 hours of treatment using the combination drugs and zoledronic acid. TGF-ß1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group and zoledronic acid group were significantly reduced compared with the control group (P<0.01). Furthermore, TGF-ß1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group were significantly lower than those of the weak combination group (P<0.01). CONCLUSION: Combination treatment of osthole, psoralen with aconitine can inhibit cancer cell invasion, which is a result of alteration of the TGF-ß/Smad signaling pathway and down-regulation of NF-κB and RANK expressions.

Hesperidin upregulates heme oxygenase-1 to attenuate hydrogen peroxide-induced cell damage in hepatic L02 cells.

Hesperidin, a naturally occurring flavonoid presents in fruits and vegetables, has been reported to exert a wide range of pharmacological effects that include antioxidant, anti-inflammatory, antihypercholesterolemic, and anticarcinogenic actions. However, the cytoprotection and mechanism of hesperidin to neutralize oxidative stress in human hepatic L02 cells remain unclear. In this work, we assessed the capability of hesperidin to attenuate hydrogen peroxide (H(2)O(2))-induced cell damage by augmenting the cellular antioxidant defense. Real-time quantitative polymerase chain reaction, Western blot, and enzyme activity assay demonstrated that hesperidin upregulated heme oxygenase-1 (HO-1) expression to protect hepatocytes against oxidative stress. In addition, hesperidin also promoted nuclear translocation of nuclear factor erythroid 2-related factor (Nrf2). What's more, hesperidin exhibited activation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). Besides, ERK1/2 inhibitor significantly inhibited hesperidin-mediated HO-1 upregulation and Nrf2 nuclear translocation. Taken together, the above findings suggested that hesperidin augmented cellular antioxidant defense capacity through the induction of HO-1 via ERK/Nrf2 signaling. Therefore, hesperidin has potential as a therapeutic agent in the treatment of oxidative stress-related hepatocyte injury and liver dysfunctions.