Scutellarin suppresses the metastasis of triple-negative breast cancer via targeting TNFα/TNFR2-RUNX1-triggered G-CSF expression in endothelial cells.

Triple-negative breast cancer (TNBC) is heterogeneous and aggressive, with high vascularity and frequent metastasis. We have already found natural flavonoid scutellarin (SC) suppressed spontaneous TNBC metastasis via normalizing tumor vasculature in vivo. In this study, supernatant from tumor necrosis factorα (TNFα)-treated human mammary microvascular endothelial cell (HMMEC) promoted cell migration and pseudopod formation in TNBC cells, but these phenomena were disappeared in SC-co-treated HMMEC. TNFα enhanced the expression of granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) in both HMMEC and human umbilical vein endothelial cell (HUVEC). G-CSF promoted TNBC migration and invasion in vitro, while G-CSF neutralization antibody and SC both inhibited TNBC metastasis in Balb/c mice. SC had no inhibition on the G-CSF-induced TNBC cell migration, but reduced G-CSF content in TNBC tumor tissues and TNFα-stimulated endothelial cells (ECs). SC restricted the nuclear translocation of runt-related transcription factor 1 (RUNX1) in TNBC tumor vessels and TNFα-treated ECs. RUNX1 was found to directly bind to the promoter of G-CSF in TNBC tumor vessels and regulated G-CSF expression. TNF receptor 2 (TNFR2) was crucial for regulating the TNFα-induced RUNX1 activation and G-CSF expression. Notably, SC hindered the interaction between TNFα and TNFR2 via binding to TNFR2. This work demonstrated that SC reduced TNBC metastasis by targeting TNFα/TNFR2-initiated RUNX1 activation and subsequent G-CSF production in TNBC-associated ECs.

Integrating network pharmacology and in vivo experimental validation to reveal the alleviation of mailuoning oral liquid on non-alcoholic fatty liver disease.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) especially the later stage non-alcoholic steatohepatitis (NASH) seriously endangers human's health and has become a global public health issue in recent years. Mailuoning Oral Liquid (MLN) is a modern traditional Chinese medicine prescription composed by Lonicerae japonicae flos, Achyranthis bidentatae radix, Scrophulariae radix and Dendrobium Caulis. MLN is generally used to treat the syndrome of blood stasis in clinical practice. PURPOSE: To observe the alleviation of MLN on NASH in vivo, and explore the possible underlying mechanism. Furthermore, this study also aims to find which Chinese medicinal drug contained in MLN exerts the main pharmacological activity. METHODS: NASH model was induced in mice by feeding with methionine and choline deficient (MCD) diet. The effects of MLN on hepatic lipids accumulation, liver inflammation, hepatic fibrosis, and the expression of some molecules were investigated by histological observation, biochemical index analysis, quantitative real-time PCR and western blot. Network pharmacology was applied to predict those involved molecular targets and potential mechanisms, which was further validated in vivo. BODIPY fluorescence staining assay was used to detect cellular lipids accumulation. RESULTS: MLN (7.8, 23.4 ml/kg) improved NASH in MCD-fed mice. Network pharmacology results demonstrated that peroxisome proliferator-activated receptor α (PPARα) signaling pathway was crucially involved in the MLN-provided alleviation on NASH. Further experimental validation results showed that MLN increased the expression of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) and restored the decreased expression of nuclear PPARα in MCD-fed mice. Further results displayed that Achyranthis bidentatae radix and Lonicerae japonicae flos contributed greatly to the MLN-provided alleviation on NASH in vivo. BODIPY fluorescence staining assay showed that 25R-inokosterone and cynaroside, two compounds from Achyranthis bidentatae radix and Lonicerae japonicae flos, obviously reduced intracellular lipids accumulation in hepatocytes stimulated by non-esterified fatty acid (NEFA). CONCLUSION: MLN improved NASH in MCD-fed mice, and the PGC-1α-PPARα signaling pathway was involved in this process. Moreover, Lonicerae japonicae flos and Achyranthis bidentatae radix contained in MLN contributed greatly to the MLN-provided improvement on NASH.

Scutellarin suppresses triple-negative breast cancer metastasis by inhibiting TNFα-induced vascular endothelial barrier breakdown.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with high vascularity and frequent metastasis. Tumor-associated abnormal vasculature was reported to accelerate TNBC metastasis. Scutellarin (SC) is a natural flavonoid with a cardiovascular protective function. In this study, SC reduced TNBC metastasis and alleviated tumor-associated vascular endothelial barrier injury in vivo. SC rescued the tumor necrosis factor-α (TNFα)-induced diminishment of endothelial junctional proteins and dysfunction of the endothelial barrier in vitro. SC reduced the increased transendothelial migration of TNBC cells through a monolayer composed of TNFα-stimulated human mammary microvascular endothelial cells (HMMECs) or human umbilical vein endothelial cells (HUVECs). TNFα induced the nuclear translocation of enhancer of zeste homolog-2 (EZH2), and its chemical inhibitor GSK126 blocked TNFα-induced endothelial barrier disruption and subsequent TNBC transendothelial migration. TNF receptor 2 (TNFR2) is the main receptor by which TNFα regulates endothelial barrier breakdown. Extracellular signal-regulated protein kinase (ERK)1/2 was found to be downstream of TNFα/TNFR2 and upstream of EZH2. Additionally, SC abrogated the TNFR2-ERK1/2-EZH2 signaling axis both in vivo and in vitro. Our results suggest that SC reduced TNBC metastasis by suppressing TNFα-initiated vascular endothelial barrier breakdown through rescuing the reduced expression of junctional proteins by regulating the TNFR2-ERK1/2-EZH2 signaling pathway.

Si-Miao-Yong-An Decoction for Diabetic Retinopathy: A Combined Network Pharmacological and In Vivo Approach.

Si-Miao-Yong-An decoction (SMYAD), a traditional Chinese medicine formula, is mainly used to clear away heat and detoxify and to promote blood circulation and relieve pain. Diabetic retinopathy (DR) is the most common type of microvascular complication caused by diabetes. This study is designed to examine the protective effect of SMYAD against DR and further to reveal the engaged mechanism via integrating network pharmacology and in vivo experimental evidence. Streptozotocin (STZ) was intraperitoneally injected into mice to induce diabetes. The dysfunction of the blood retina barrier (BRB) was observed by conducting Evan's blue leakage assay, detecting tight junction (TJ) protein expression and counting the number of acellular capillaries in retinas. Our results showed that SMYAD alleviated BRB breakdown in vivo. Network pharmacology results demonstrated that regulating inflammation, immune responses, and angiogenesis might be associated with the efficacy of SMYAD in alleviating DR, in which the tumor necrosis factor (TNF) and hypoxia inducible factor 1 (HIF1) signal pathways were involved. Next, immunofluorescence staining results showed that SMYAD decreased microglia activation in retinas and reduced the enhanced adhesion of leukocytes into retinal vessels. SMYAD reduced the elevated serum TNFα content and retinal TNFα expression. SMYAD abrogated the activation of nuclear factor κB (NFκB) and HIF1α and consequently decreased the enhanced expression of some pro-inflammatory molecules and vascular endothelial growth factor (VEGF) in retinas. These results indicate that SMYAD attenuated DR development through suppressing retinal inflammation and angiogenesis via abrogating NFκB-TNFα and HIF1α-VEGF signal pathways.